Stem Cell Transplantation _Cord Blood Transplants_ by pengxiuhui


									       Stem Cell Transplantation (Cord Blood Transplants)
                   Nelson J. Chao, Stephen G. Emerson, and Kenneth I. Weinberg

Allogeneic stem cell transplantation is an ac-              In Section II, Dr. Stephen Emerson describes
cepted treatment modality for selected malignant       the historical efforts associated with expansion
and non-malignant diseases. However, the ability       of hematopoietic stem cells, specifically with
to identify suitably matched related or unrelated      cord blood cells. These efforts to expand cord
donors can be difficult in some patients. Alterna-     blood cells continue with novel methods. More-
tive sources of stem cells such as cord blood          over, a better understanding of stem cell biology
provide a readily available graft for such patients.   and signaling is critical if we are to be able to
Data accumulated over the past several years           effectively expand these cells for clinical use. An
have demonstrated that the use of cord blood is        alternative, more direct, approach to expanding
an accepted source of stem cells for pediatric         stem cells could be achieved by specific genetic
patients. Since the cell numbers of hematopoi-         pathways known or believed to support primitive
etic progenitors in cord blood is limited and the      HSC proliferation such as Notch-1 receptor
collection can occur only in a single occasion,        activation, Wnt/LEF-1 pathway induction,
its use in adult patients can be more problem-         telomerase or the Homeobox (Hox) gene prod-
atic. Here, new developments in the use of cord        ucts. The clinical experience with the use of
blood for adults and studies aimed at expansion        expanded cord blood cells is also discussed.
of cord blood cells and immune reconstitution               In Section III, Dr. Kenneth Weinberg de-
are described.                                         scribes immune reconstitution or lack thereof
     In Section I, Dr. Nelson Chao describes the       following cord blood transplantation. One of the
early data in cord blood transplantation in adult      hallmarks of successful hematopoietic stem cell
patients. The patient outcomes are reviewed and        transplantation is the ability to fully reconstitute
analyzed for various factors such as cell dose,        the immune system of the recipient. Thus, the
HLA typing, and patient selection that could have      relationship between stem cell source and the
contributed to the final outcome of these adult        development of T lymphocyte functions required
patients. Myeloablative as well as nonmyelo-           for protection of the recipient from infection will
ablative approaches are presented. Discussion of       be described, and cord blood recipients will be
the various benefits and risks are presented.          compared with those receiving other sources of
More recent data from multiple single institu-         stem cells. T cell development is described in
tions as well as larger registry data comparisons      detail, tracking from prethymic to postthymic
are also provided. Analyses of these studies           lymphocytes with specific attention to umbilical
suggest methods to improve on the outcome.             cord blood as the source of stem cells. Moreover,
These newer data should lead to a logical              a discussion of the placenta as a special micro-
progression in the use of cord blood cells in          environment for umbilical cord blood is pre-
adult patients.                                        sented. Strategies to overcome the immunologi-
                                                       cal defects are presented to improve the outcome
                                                       of these recipients.

354                                                                       American Society of Hematology
           I. CORD BLOOD TRANSPLANTS:                         eages and the enrollment of high-risk patients with ad-
 HOW    CLOSE ARE WE TO USING THIS IN ADULTS?                 vanced disease all contributed to this caution. Recently,
                                                              older allogeneic recipients have been treated success-
                  Nelson J. Chao, MD*                         fully following a variety of less intense nonmyelo-
                                                              ablative conditioning regimens. On the basis of these
Umbilical cord blood transplantation (UCBT) has re-           encouraging observations, it has been hypothesized that
cently been explored in an increasing number of adult         a reduced-intensity preparative regimen would allow
patients. The relative ease of procurement and the lower-     engraftment of the UCB stem cells. While the total num-
than-anticipated risk of severe acute graft-versus-host       bers of mononuclear cells are limited, the progenitor
disease (GVHD) has made UCBT an appealing alterna-            content and the proliferative potential of cord blood
tive to bone-marrow–derived hematopoietic stem cells.         cells are high. This nonmyeloablative approach could
The use of reduced-intensity or nonmyeloablative pre-         diminish some of those concerns raised with the use of
parative regimens to allow engraftment of UCBT broad-         ablative regimens. This review will summarize the avail-
ens the scope of patients who may benefit, including          able data on the use of UCB as an alternative source of
older and medically infirm patients without a matched         hematopoietic stem cells for allogeneic transplantation
donor. This section will summarize the available data         for adult patients.
on the use of UCB as alternative source of hematopoi-
etic stem cell transplantation in adult patients.             Myeloablative Preparative Regimens
     The first known attempt at UCBT occurred 34 years        In comparison to the published studies on pediatric pa-
ago when a young 16-year-old boy with acute lympho-           tients, the clinical data on the use of UCBT in adult
blastic leukemia received cord blood units from 8 dif-        patients are relatively limited, although growing7-14
ferent donors. He was receiving 6-mercaptopurine and          (Table 1). As can be gleaned from the composite data,
prednisone and did not receive a preparatory regimen.         these patients received standard total body irradiation
One unit engrafted as demonstrated by red cell antigens       or busulfan-based myeloablative regimens. GVHD pro-
and the graft lasted for 38 days.1 However, it was the        phylaxis consisted of cyclosporine alone or in combi-
success of the allogeneic UCBT approximately 16 years         nation with steroids or methotrexate. The degree of HLA
ago in a child with Fanconi anemia that opened a new          disparity was not associated with outcome in any of
source of allogeneic hematopoietic stem cells.2 UCB is        these studies, although the numbers in each were small.
now accepted as an alternative source for hematopoi-          However, in a recent review update with 861 unrelated
etic stem cells for transplantation in children. Unre-        UCBT recipients from the Placental Blood program of
lated UCB offers many practical advantages as an alter-       the New York Blood Center, which includes 181 (21%)
native source of stem cells, including: (1) the relative      patients with age ≥ 18 years and 170 patients (20%)
ease of procurement and availability (given the ability       weighing ≥ 60 kg, Rubinstein et al15 have demonstrated
to store fully tested and HLA-typed cord blood avail-         in a multivariate analysis that HLA match is an inde-
able for immediate use); (2) the absence of risk for          pendent predictor of event-free survival.
mothers and donors; (3) the reduced likelihood of trans-           The median age varied considerably from a low of
mitting infections, particularly cytomegalovirus; (4) po-     29 to a high of 48 years with a range of 15 to 58 years.
tential reduced risk of GVHD; (5) less stringent criteria     Age is an important factor in the outcome, with the
for HLA matching for donor-recipient selection (with          younger patients generally doing better than the older
the potential of finding donors for minority popula-          patients. The median weight for these studies also var-
tions); and (6) absence of donor attrition. UCB banks         ied from a low of 51 to 70 kg. As the cell dose of the
have been established for related and unrelated UCBT          UCB is given per recipient weight, the patients with
with about 100,000 units currently available.3-6              lower body weights would have a higher chance of re-
     While the clinical data are encouraging for pediat-      ceiving a higher cell dose per cord blood unit. The range
ric patients, the limited number of hematopoietic stem        of mononuclear cells/kg recipient weight ranged from
cells in UCB has been a cause for caution for its use in      1.5 to 2.43 × 107/kg. The total numbers of CD34+ cells
adult patients. Moreover, the use of myeloablative pre-       has been correlated with both engraftment and the speed
paratory regimens with their known toxicities together        of engraftment.
with the known delays in engraftment of all cell lin-              The diseases for which these patients were trans-
                                                              planted consisted primarily of hematological malignan-
                                                              cies. The differences in outcome can be partially attrib-
* Duke University, 2400 Pratt Street, Suite 1100, Durham NC   uted to the proportion with more favorable disease sta-
27705-3976                                                    tus at transplantation. For example, most of the data

Hematology 2004                                                                                                   355

                                 Table 1. Studies of unrelated umbilical cord blood (UCB) transplantation in adults.

                                                                                                                           Rocha et al7,9                                Cornetta et al11
                                                                           Laughlin et al7           Sanz et al8            (Eurocord)           Goldberg et al10           (COBLT)           Ooi et al12            Ooi et al13               Long et al14
                                 Number of patients                              68                      22                     108                     19                      34                18                      13                        57*
                                 Median Age (range, years)                31.4 (17.6–58.1)           29 (18–46)              26 (15–53)             48 (20–59)            34.5 (18.2–55)      43 (21–52)             40 (20–51)                 31 (18–58)
                                 Median weight (range, kg)               69.2 (40.9–115.5)           70 (41–85)             60 (35–110)             69 (52–126)                 NA             55.2 (NA)             51 (43–68)                70 (46–110)
                                 Diseases (n)
                                   Hematological malignancies                    54                      21                      96                     18                      32                18                      13                        50
                                   Bone marrow failure syndrome/MDS              13                       1                      12                      1                       2                                                                   6
                                 Number of HLA loci disparity (n)
                                   0                                              2                       1                       6                      4                       1                 0                      0                          2
                                   1                                             18                      13                      38                      8                      10                 4                      4                          8
                                   ≥ 2                                           48                       8                      64                      7                      23                14                      9                         47
                                 Preparative regimen (n)                  TBI-based/ATG           Thio/Bu/Cy/ATG                NA                  Cy/TBI/ATG                Cy/TBI         TBI/Ara-C/Cy           Cy/TBI/Ara-C              Cy or Mel with
                                                                              (n = 51)                (n = 21)                                       (n = 13)                (n = 27)          (n = 17)               (n = 13)                  TBI or BU/
                                                                           Bu-based/ATG            Thio/Flu/ATG                                    Mel/TBI/ATG                Bu/Mel         TBI/Ara-C/Flu                                         ATG
                                                                              (n = 14)                 (n = 1)                                        (n = 4)                 (n = 7)           (n = 1)                                           n = 41
                                                                         Others/ATG (n = 4)                                                        Bu/Cy (n = 2)                                                                            TAI/Cy/ATG (n = 1)
                                 GVHD prophylaxis                        CSA/Pred (n = NA)       CSA/Pred (n = 22)       CSA/Pred (n = 77)      CSA/Pred (n = 13)            NA              CSA (n = 2)         CSA/ MTX (n = 13)            CSA (n = 1)
                                                                           CSA (n = NA)                                   Others (n = 31)       FK506/Pred (n = 6)      CSA/MTX (n = 16)                                                    CSA/Pred (n = 56)
                                 Median time to engraftment (range,days)
                                   ANC > 500/µLµ                         27 (13–59)                  22 (13–52)             32 (13–60)                28 (NA)              28.5 (13–55)      23 (16–41)              22.5 (19–35)               26 (12–55)
                                   Platelet > 20,000/µL                  58 (35–142)                69 (49–153)            129 (26–176)               56 (NA)                   NA          49 (31–263)**           49 (30–164)**              84 (35–167)
                                 Probability of myeloid engraftment       90% by 42 days          100% at 60 days         81% by day 60           75% by day 60           72% at day 60          94%                     92%                  80% at day 50
                                 GVHD (n) ( probability, %)
                                   Acute Grade II-IV                          33 (60%)                16 (NA)                 44 (38)                   NA                   11 (38%)          10 (55%)                 9****                    17 (41%)
                                   Acute Grade III-IV                         11 (20%)                7 (NA)                  27 (NA)                   NA                    6 (21%)           1 (6%)                    –                       9 (22%)
                                   Chronic/ patients at risk                   12/33                   9/10                    15/58                    NA                       NA             14/18                    8/11                       8/25
                                 Median cell dose (range)
                                   N.C. infused (x 107/kg)                   1.6 (0.6–4)          1.71 (1.01–4.96)          1.71(0.2–6)            1.8 (0.4–5.3)         1.73 (1.11–3.75) 2.51 (1.16–5.29)         2.43 (2.09–4.06)           1.5 (0.54–2.78)
                                   CD34+ cells infused (x 105/kg)           1.2 (0.2–16.7)        0.79 (0.27–2.60)              NA                      NA                      NA               NA                       NA                 1.37 (0.02–12.45)
                                 Therapy-related mortality (%)            50% at 100 day          43% at 100 day          54% at 100 day                NA                      NA               6%                      0%                        56%
American Society of Hematology

                                 Survival (%)                                    NA                      NA                27% at 1 year         20% for good risk       30% at 180 days 76% at 2 years           76.2% at 2 years            19% at 3 years
                                                                                                                                               and 21% for poor risk
                                                                                                                                                    at 1 year
                                 Event-free survival (%)                26% at 40 monthsa          53% at 1yearb          21% at 1 yearc                NAd                    NAe          77% at 2 yearsf              NAg                 15% at 3 yearsh
                                 Findings and Comments:
                                 a EFS is better in patients receiving graft with CD34+ cells >1.2 x 105/kg. b Patients under the age of 30 had significantly better survival. c100 day TRM is lower in patients with disease in chronic phase or remission,
                                 N.C.dose infused ≥ 2.0 x 107/kg and transplant performed after January 1998. d 10 patients received ex vivo expanded UCBT. e The high mortality and relapse reflects the poor risk patients enrolled. . f All de novo AML.
                                 g All except 1 patients received UCB transplant as an upfront treatment and all patients received > 2 x 107 N.C. per weight, perhaps due to smaller size of patients (median weight 51 kg). h CD34+ cell dose infused
                                 correlated with rate of platelet recovery and age > 31 years was significant predictor of poorer event-free survival.
                                 *19 of the 57 patients were included in the study published by Laughlin et al.7
                                 ** platelet > 50,000/µL
                                 *** All had MDS-related secondary acute myeloid leukemia
                                 **** 9 of 12 evaluable patients had acute acute graft-versus-host disease.
                                 Abbreviations: NA, not available; TRM, therapy-related mortality; N.C., nucleated cell dose; TAI , thoraco-abdominal radiation; Ara-C, cytarabine arabinoside; TBI, total body irradiation; ATG, antithymocyte globulin;
                                 Bu, bulsulfan; Cy, cyclophosphamide; Thio, thiotepa; Flu, fludarabine; Mel, melphalan; Pred, prednisone; CSA, MTX, methotrexate; GVHD, graft-versus-host disease; MDS, myelodysplastic syndrome
from Sanz et al8 or Ooi et al13 included recipients with        appears that a UCB graft that contains at least 2 × 107
chronic myeloid leukemia (CML) in first chronic phase           nucleated cells/kg and 1.7 × 105 CD34+ cells/kg is ac-
or acute leukemia in first remission or de novo acute           ceptable for adult recipients.4-6 It is hoped that the ad-
myeloid leukemia or untreated myelodysplastic syn-              vantage of a lower GVHD without any apparent in-
drome. In contrast, the less favorable survival in the          crease in relapse will offset the adverse impact of re-
other studies reflects the poor-risk subjects enrolled into     duced cell dose on survival. As in the pediatric setting,
the study.                                                      TRM remains the main obstacle for success in adult
     The probability of myeloid engraftment ranged from         recipients, although several investigators have demon-
72% to 100% by day +60; however, some patients were             strated that the TRM is not different from that seen
censored to this event if they died prior to the set date       with matched unrelated donors. With the profound in-
for the definition of engraftment. The median times to          fluence of UCB cell dose (both nucleated cell dose and
engraftment of both neutrophils and platelets varied as         CD34+ cell dose) on engraftment, survival and prob-
well with approximately 1 month to reach 500 neutro-            ably TRM in the adult setting, future research should
phils/mL and 2–4 months to reach platelet counts                also focus on increasing the cell dose of the UCB graft.
> 20,000/µL. Again, the pace of recovery is likely to           Possible methods to increase cell dose include ex vivo
reflect the UCB stem cell dose as well as the amount of         expansion, infusion of multiple cord blood units or use
prior therapies a patient could have received. As com-          of possible additional cells such as mesenchymal stem
pared to the pediatric patients, the incidence of severe        cells or improving HLA matching.
grade III-IV acute GVHD was low (approximately 20%)
given that the majority of the patients received HLA            Nonmyeloablative Regimens
mismatched unrelated grafts. Of note, the HLA match-            The nonmyeloablative stem cell transplantation (NST)
ing in general was done serologically for HLA-A and-            regimen was proposed initially based on the rationale
B and molecularly for HLA-DR. Thus it is likely that            that the therapeutic benefit of an allogeneic transplan-
the mismatching would have been greater than reported           tation is partially related to the crucial immune-medi-
if molecular matching had been performed. Therapy-              ated graft-versus-malignancy effect. The concept of
related mortality (TRM) was high, reaching approxi-             graft-versus-malignancy as the pivotal therapeutic com-
mately 50% in most series. The overall survival again           ponent of allogeneic transplantation is supported by the
varied considerably, reflecting the risk factors outlined       observation from clinical studies that (1) patients with
above. Comparative studies of UCB transplantation               acute and chronic GVHD have a reduced risk of re-
versus unrelated bone marrow have been performed from           lapse; (2) patients with syngeneic BMT and after T cell–
two registry datasets. These two studies have demon-            depleted allotransplant have a higher incidence of re-
strated that if there is a cord blood unit that is at least a   lapse compared with other allogeneic donors; and (3)
4/6 HLA match with sufficient cells, the outcomes are           patients with a relapsed malignancy after allogeneic
similar following UCB transplantation to matched or             transplantation can be re-induced into complete remis-
one antigen mismatched unrelated donor bone marrow              sion without any chemotherapy by donor lymphocyte
grafts.21,22                                                    infusion (DLI). NST, with its reduced-intensity pre-
     Thus, UCB contains sufficient numbers of hemato-           parative regimen, makes allogeneic transplantation ap-
poietic stem cells to achieve engraftment in adult pa-          plicable to patients with relative contraindications to
tients with lower than anticipated risk of severe acute         myeloablative regimens given its lower rate of TRM
GVHD, even when HLA-disparate grafts are infused.               with fewer infections and less GVHD. However, the
The use of UCB allows allografting to be offered to a           preparative regimen does not contribute significantly
greater proportion of patients, many of whom do not             to the antimalignancy effect; thus, the risk of disease
have a matched sibling or unrelated donor to allow al-          progression after transplantation remains higher com-
logeneic therapy as the only chance to cure the under-          pared with the myeloablative approach.
lying disease. The results thus far suggest that UCBT                NST using UCB provides an opportunity for im-
can result in long-term disease-free survival in many of        munotherapy for older patients, sicker patients and also
these patients. Similar to the pediatric series, clinical       patients without suitable donors who are not eligible
experience in the adult patients has also documented            for this potentially curative approach. However, there
the importance of graft cell dose in determining en-            is increased concern about graft rejection using this ap-
graftment and survival. The critical “threshold dose,”          proach given that there are, on average, two logs fewer
below which engraftment and survival become signifi-            cells infused than would be considered for a standard
cantly inferior, remains to be defined in a larger study        matched sibling or unrelated donor transplant (Table
with a longer follow-up. Based on current results, it           2). The clinical outcome of 2 patients with malignant

Hematology 2004                                                                                                      357
Table 2. Summary of results of nonmyeloablative unrelated umbilical cord blood transplantation in adult patients.

Investigator                                Barker et al18                McSweeney et al19                 Chao et al20
Number of patients                                43                                  5                          13
Age, years: median (range)                   49.5 (22–65)                    64.5 (25–78)                    49 (19–62)
Diagnosis                                         HM                      HD; AML; CLL; NHL           NHL; MDS; AML; MEL; ALL
Preparative regimen                 F + Bu + TB I (200cGy (n = 21)         F + TBI (200cGy)             F + C + ATG (n = 10)
                                    F + C + TBI (200cGy) (n = 22)                               F + C + ATG + TBI (200cGy) (n = 3)
GVHD prophylaxis                              CYA + MMF                       CYA + MMF                  CYA + Pred (n = 8)
                                                                                                         CYA + MMF (n = 5)
Cell Dose
N.C. (× 107/kg): median (range)              2.6 (1.6–3.8)†                  1.1 (1.75–1.3)               2.07 (1.07–5.53)
                                             3.2 (1.1–5.1)‡
CD34 (× 106/kg): median (range)             3.7 (1.1–8.1)†                  NA (0.01–0.04)                  1.3 (0.5–9.6)
                                            4.3 (1.1–10.3)‡
CD3 (× 106/kg): median (range)             0.06 (0.02–0.15)†                 NA (1.4–3.3)                 4.6 (2.02–22.82)
                                           0.05 (0.02–0.12)‡
Engraftment (n)                               C.I. = 76%†                             2                         8/12
                                              C.I. = 94%‡
Median days to ANC                            26 (12–30)†                         NA                         12 (6–24)
> 500/µL (range)                              9.5 (5–28)‡
Median days to platelet                                                           NA                         14 (6–61)
> 20,000/µL (range)
Grade II-IV aGVHD / cGVHD (n)           C.I. for aGVHD = 44%                     1/NA                            1/1
                                        C.I. for cGVHD = 21%
Outcome                                  O.S. : 39% at 1 year                 Alive: n = 3               O.S.: 43% at 1 year
                                         DFS.: 31% at 1 year                   PR: n = 1                 DFS: 43% at 1 year

Abbreviations: Gd, grade; aGVHD, acute graft-versus-host disease; cGVHD, chronic graft-versus-host disease; N.C., nucleated
cell; ANC, absolute neutrophil count; NHL, non-Hodgkin’s disease; MDS, myelodysplastic syndrome; AML, acute myeloid leukemia;
MEL, melanoma; ALL, acute lymphoblastic leukemia; HD, Hodgkin’s disease; CLL, chronic lymphocytic leukemia; THAL, thalas-
semia; WAS, Wiskott–Aldrich syndrome; NBL, neuroblastoma; HM, hematological malignancies; F, fludarabine; ATG, antithymocyte
globulin; C, cyclophosphamide; TBI, total body irradiation; Bu, busulphan; CYA, cyclosporin A; MMF, mycophenolate mofetil; PDN,
prednisolone; FK506, tracrolimus; NA, not available; C.I., cumulative incidence; PR, partial remission; OS, overall survival; DFS,
disease-free survival
†   The results refer to patients given F + Bu + TBI 200cGy as conditioning regimen.
‡   The results refer to patients given F + C + TBI 200cGy as conditioning regimen.

lymphoma using this novel approach was first reported                potential for robust engraftment using this approach.
by investigators at Duke University.16,17 In that study, 2           The favorable outcome demonstrates the feasibility of
patients with relapsed lymphoma who had no matched                   the mismatched unrelated UCB cells, even with the NST
siblings, partially matched family members, or matched               regimens.
unrelated donors successfully underwent nonmyelo-                         The largest experience with transplantation using a
ablative conditioning therapy followed by infusion of                reduced-intensity regimen was evaluated by investiga-
4 and 6 matched, unrelated donor UCB cells, respec-                  tors from the University of Minnesota on a cohort of
tively, at the nucleated cell dose of 2.9 and 6.5 × 107/             high-risk patients with hematological malignancies.18
kg, respectively. The conditioning regimens consisted                In their study, unrelated UCB graft with a median nucle-
of fludarabine 30mg/m2 and cyclophosphamide 500 mg/                  ated cell dose of 3.7 × 107 per kg (range, 1.6–6.0 × 107
m2 daily for 4 days with antithymocyte globulin 30                   /kg) was infused into 43 patients (median age 49.5 years;
mg/kg per day for 3 days. Cyclosporine and predniso-                 range, 22–65 years), after receiving 2 types of condi-
lone were given for acute GVHD prophylaxis. Both                     tioning regimens: fludarabine 200 mg/m2, total body
patients had 100% donor engraftment by the third month               irradiation (TBI) 200 cGy and busulfan 8 mg/kg (Flu/
of transplant and remained in remission 6 and 12 months              Bu/TBI) for the initial 21 subjects; fludarabine 200 mg/
following transplantation. Extension of these data to                m2, TBI 200 cGy and cyclophosphamide 50 mg/kg (Flu/
11 other patients confirms the low morbidity and the                 Cy/TBI) for the subsequent 22 subjects. All patients

358                                                                                           American Society of Hematology
received GVHD prophylaxis with cyclosporine and              able to expand effectively in the periphery, and the de-
mycophenolate mofetil. The median time to neutrophil         velopment of new T cells through the thymus is also
recovery of more than 0.5 × 109/L was 26 days (range,        accelerated compared to the rate of development in those
12–30 days) for the Flu/Bu/TBI recipients, but only          receiving ablative regimens. Alternatively, the lower
9.5 days (range, 5–28 days) for the Flu/Cy/TBI recipi-       incidence of GVHD in NST may also play an impor-
ents. The cumulative incidence of engraftment for Flu/       tant role in the preservation of the peripheral and cen-
Bu/TBI and Flu/Cy/TBI recipients was 76% and 94%,            tral niches for T cell development.
respectively. Despite the use of 1–2 HLA-antigen mis-
matched graft in 93% of the recipients, the cumulative       Future Directions for UCB Transplantation
incidence of grade II–IV GVHD and grade III–IV                   in Adults
GVHD for the entire cohort of patients were 44% and          While the use of NST signifies an advancement in the
9%, respectively. The disease-free survival of these high-   field of transplantation and immunobiolology, several
risk subjects was also favorable: 24% at 1 year for Flu/     unresolved questions remain:
Bu/TBI recipients and 41% at 1 year for Flu/Cy/TBI            1. What is the optimal UCB cell dose required for
recipients. A similar approach and data have also been            adult patients and can we increase it?
reported by investigators in University of Colorado           2. What is the optimal NST conditioning regimen and
Health Sciences Center.19                                         GVHD prophylaxis?
     Immune recovery in 5 recipients of UCB trans-            3. Is the incidence of GVHD similar in ablative and
plant following the NST regimen were compared to                  NST regimens?
recovery in adult recipients of UCB following a myelo-        4. Does the incidence of infection differ in ablative
ablative regimen by investigators at Duke University.20           versus NST regimens and can one improve immune
Recipients of NST regimens had a more rapid and ro-               recovery?
bust recovery of myeloid and lymphoid cells. The T            5. What is the difference in overall efficacy between
cell repertoire in UCB recipients treated with the NST            myeloablative and NST UCBT?
regimen was markedly more diverse and robust com-
pared with the repertoire in those receiving the             Conclusion
myeloablative regimen at similar time points. T cell         UCB is a viable alternative to bone marrow and periph-
receptor excision circles (TRECs), which are generated       eral blood as a source of stem cells capable of hemato-
within the thymus and identify new thymic emigrants          poietic reconstitution for adults, when related or unre-
and those that have not divided, were detected 12 months     lated marrow donor is not available. UCBT following
after transplantation in the NST recipients. This com-       NST preparative regimen is an exciting new approach
pared favorably to the delayed detection of TRECs at         that provides an option for patients who are otherwise
18–24 months among recipients of myeloablative regi-         excluded from conventional hematopoietic stem cell
mens. Thus, in adults receiving an NST preparatory           transplantation, including elderly or medically infirm
regimen, the quantitative and qualitative recovery of T      patients with no matched sibling donor. Preliminary
cells occurs through rapid peripheral expansion. The         results have shown that such an approach can be associ-
ability of patients receiving NST transplantation to re-     ated with timely engraftment with full donor chimer-
cover within a few months suggests that the peripheral       ism. Comparison between myeloablative and NST ap-
“niches” in which T cells can proliferate are preserved      proaches will be needed before this therapy can be con-
in these patients compared to those receiving                sidered for younger patients eligible for myeloablative
myeloablative regimens. Moreover, the presence of            transplant. At the moment, the use of NST UCBT can-
TREC-positive cells within 1 year suggests that thymic       not be encouraged outside of a clinical trials or selected
recovery is likewise accelerated in NST compared with        patients. The future challenge will be to develop strat-
recipients of myeloablative regimens. The favorable          egies to optimize the chance of early and durable en-
results of T cell recovery following the NST suggest         graftment, as well as to minimize the risk of GVHD
that it may be possible to have an excellent outcome         and transplant-related death.
with an unrelated mismatched UCB transplantation in
adult patients. Patients have a rapid recovery of T cells
with a complex diversity. The primary difference be-
tween the recipients of ablative and NST regimens was
the extent of physiologic damage caused by the prepa-
ratory regimen. When the damage is relatively mild, as
in nonmyeloablative regimens, the donor T cells are

Hematology 2004                                                                                                   359
  II. EXPANSION OF UMBILICAL CORD BLOOD CELL                   shorten UCB transplant nadirs by providing increased
   PROGENITORS AND STEM CELLS: BIOLOGY AND                     numbers of stem and progenitor cells to patients by
       APPLICATION TO CLINICAL TRANSPLANTS                     expanding the numbers of hematopoietic stem cells
                                                               (HSCs) and progenitor cells, in the laboratory, prior to
            Stephen G. Emerson, MD, PhD*                       transplant. This approach should succeed if the delay in
                                                               engraftment is due to a paucity of HSC and progenitor
The close correlation between preclinical assays and in        numbers, but would fail if the delay were due to an
vivo clinical biology that is observed with bone mar-          uncharacterized-but-insurmountable qualitative defect
row stem cells and with hematopoietic growth factors           in UCB stem and progenitor cells.
such as erythropoietin (Epo), granulocyte colony-stimu-
lating factor (G-CSF) and granulocyte-macrophage               Stem Cells, Stem Cell Assays, and Engraftment
colony-stimulating factor (GM-CSF) has been very re-           Attempts to characterize UCB expansion protocols have
assuring and has accelerated the development and im-           relied directly on the assays used to measure the cell
provement of clinical hematology practice. Clinical ex-        populations that are targets for expansion. Over the past
perience with UCB transplants, in contrast, has proved         20 years, these assays have become more sophisticated,
much more paradoxical and problematic. Early labora-           and clinical investigators have progressively adjusted
tory measurements showed that the density of both              their clinical trial designs based on more recent, better
CD34+ cells and clonogenic progenitor cells was ex-            assays. Pluripotent hematopoietic stem cells are very
tremely high in UCB.1 Similarly, the proliferative ca-         rare, representing only 1 cell in 2–10 × 106 BM cells,
pacity of progenitor cell differentiation in laboratory        perhaps slightly more frequent in UCB. There is no
assays and of UCB stem cells transplanted into immu-           phenotypic assay, to date, that directly identifies and
nodeficient mice was measurably higher than for pro-           measures these rare cells. Rather, all efforts to measure
genitors and stem cells from bone marrow (BM) or               HSCs rely on functional assays. The easiest and most
peripheral blood (PB).2,3 But when comparable num-             popular assays have been colony-forming cell (CFC)
bers of UCB progenitors (1–5 × 106/kg) and stem cells          assays, which measure cells that are partially differen-
are transplanted as are delivered to patients with stan-       tiated between HSCs and mature blood cells. These cells,
dard BM or PB grafts, engraftment is far slower, and           which correlate closely but inexactly with CD34+ cell
graft failure is more likely.4 Given that opportunistic        numbers, produce easily detectable blood cell colonies
infection and organ failure rise exponentially with time       in 2 weeks in the laboratory. But careful cell fraction-
to engraftment, and that overall long-term survival cor-       ation studies have shown that the intermediate-matu-
relates closely with platelet engraftment, this delay has      rity cells detected in these assays contribute only very
proved to be a major barrier to adoption of what would         slightly, and temporarily, to hematopoietic recovery after
otherwise be an overwhelmingly attractive stem cell            transplantation, since they don’t produce enough cells
source.                                                        that survive long enough after transplantation; it is only
     One of the striking lessons of BM and peripheral          the immature pluripotent stem cell population that con-
stem cell (PSC) transplant is that time to engraftment         tributes to sustained engraftment.5,6 So, although CD34+
correlates inversely with progenitor and stem cell dose,       cell numbers in an unmanipulated graft may correlate
with most graphed comparisons showing hyperbolic               with clinical time to engraftment after transplantation,
curves with asymptotic limits at 7–8 days for neutro-          that is likely because the ratio of intermediate progeni-
phil engraftment and 10–12 days for platelet engraft-          tor cells to very primitive stem cells is fairly constant
ments, both achieved with PSC stem and progenitor              between clinical samples, and therefore CD34+ cell num-
cell doses of > 5 × 106 CD34+ cells per kg. Providing          bers indirectly reflect primitive stem cell numbers trans-
lower doses of early hematopoietic cells results in in-        planted. This assumption, however, may not be true
creasingly longer nadirs, but providing higher doses           following many ex vivo expansion procedures, in which
does not further accelerate hematopoietic recovery.            progenitors may be greatly expanded but primitive stem
Based on these well-documented observations, the ques-         cells may not.7
tion has been raised whether one could significantly                Better functional assays for primitive stem cells,
                                                               developed in the late 1980s and 1990s, include the long-
                                                               term culture initiating cell (LTCIC), cobblestone area
                                                               forming cells (CAFC), and best of all the human to
* Abramson Cancer Center; Univeristy of Pennsylvania,
Departments of Medicine, Pathology, and Pediatrics, 510        immunodeficient NOD/SCID mouse (NOD/SRC) re-
Maloney Bldg., 36th & Spruce Streets, Philadelphia PA 19104-   populating cell assay.8 In evaluating UCB expansion
4283                                                           protocols, it is important to determine what popula-

360                                                                                 American Society of Hematology
tions were measured and specifically whether LTCIC,             effect reflects quantitative increases in HSCs or a quali-
CAFC or NOD/SRC were identified as being expanded.              tative improvement in their behavior is not certain.12
                                                                This may be the basis of the reported enhancement of
UCB Expansion Ex Vivo: Cytokine Cocktails                       UCB engraftment achieved by supplementing UCB
When UCB progenitors were first detected, investiga-            grafts with mesenchymal stem cells. UCB HSCs are
tors noted that the progenitor cells formed colonies in         selectively responsive to interleukin (IL)-6 stimula-
agar more rapidly than did BM or PSC progenitors,               tion,13 and recent studies suggest that combining UCB
and that a higher fraction of UCB colonies could be             CD34+ cell selection with IL-6/soluble IL-6 receptor/
replated to form secondary colonies. These data on UCB          SCF/Flt-3l/Tpo stimulation induces NOD/SRC expan-
progenitors, together with their much higher frequency          sion 4–5X. Overall, it appears that UCB stem cell ex-
in the UCB mononuclear cell fraction, suggested that            pansion may be possible with ex vivo cytokine cock-
they were more proliferative than their adult counter-          tails, but conditions that can generate such HSC expan-
parts. In the early 1990s Moore and Hoskins applied             sions, as opposed to the expansion of more mature pro-
the techniques of PB CD34+ cell isolation followed by           genitor populations, may have not yet been used in clini-
culture in high concentrations of multiple hematopoi-           cal trial settings.
etic growth factors first pioneered by Haylock et al to
UCB, with prima facie very promising results. After                 Stem Cell Expansion: Hox Gene Products
removal of more mature CD34– cells, clonogenic pro-                 Individual cytokines are known to trigger many intra-
genitors could be expanded 50- to 100-fold, while                   cellular signal transduction pathways, which presum-
slightly more primitive ∆ cells could be multiplied 10–             ably activate multiple gene transcription pathways.
20X, all in 2 weeks.9 Subsequently, many investigators              Thus, in retrospect, combinations of cytokines might
confirmed that UCB progenitor populations could be                  well be expected to have multiple effects on HSCs that
readily expanded, and some showed that LTCIC could                  might not permit UCB stem cell expansion. An alterna-
also be expanded 10X, particularly if combinations of               tive, more direct, approach to expanding stem cells
low-dose cytokines were combined with perfusion con-                might focus on the specific genetic pathways known or
ditions that simultaneously removed the secreted prod-              believed to support primitive HSC proliferation. Over
ucts of maturing blood cells and supplied continuous                the past decade several such pathways and transcription
fluxes of cytokines.10 Of note, however, no assays for              factors have been proposed, including Notch-1 recep-
primitive NOD/SRC repopulating cells were measured                  tor activation, Wnt/LEF-1 pathway induction and
in these studies, so it is not known to what extent, if             telomerase. One of the most promising intracellular tar-
any, true HSC were expanded in these protocols. When                gets for HSC expansion are Homeobox (Hox) gene
Lewis et al assayed UCB NOD/SRC repopulating cells                  products. These transcription factors, which were first
in parallel with CFC and LTCIC during ex vivo cytokine              identified as master switch transcriptional regulators of
expansion cultures, they found that while CFC were in-              early development, were discovered a decade ago to be
creased 20–25X, LTCIC were only increased 40%, and                  expressed in the most primitive HSCs. When cloned
NOD/SCR were maintained but not increased at all.11                 and overexpressed in mouse HSCs, several of these Hox
     True UCB NOD/SRC stem cell expansion may be                    genes induce HSC proliferation, in addition to more or
achievable by very judicious combinations of HSC pu-                less disruption in differentiation.
rification and cytokine selection. SDF-1 treatment in-                   One of these Hox genes, HoxB4, has proven to be
creases the engraftment of UCB HSCs, but whether this               the most interesting to date. HoxB4 is highly expressed
                                                                         in primitive HSCs, and its expression declines with
                                                                         differentiation. Overexpression of HoxB4 mRNA
Table 3. Currently available assays of hematopoietic progenitor
and stem cells in umbilical cord blood.
                                                                         by retroviral infection results in HSCs that expand
                                                                         over 100X if transplanted in very low numbers
             Ease and Rapidity for            Correlation with           (analogous to the clinical HSC scenario).14 Of even
Assay         Clinical Application         Stable Engraftment            more interest, recent experiments suggest that the
Colony assays           +/–         Indirect, unmanipulated grafts only same effect can be achieved by treating purified
CD34+                  +++          Indirect, unmanipulated grafts only HSCs with a modified form of soluble HoxB4 pro-
LTCIC                    –                      Possibly direct          tein (TAT-HoxB4), which increases intracellular
NOD/SRC                  –                       Likely direct           HoxB4 protein levels for several hours only.15
                                                                         While this approach has thus far only been applied
Abbreviations: LTCIC, long-term culture-initiating cell; NOD/SRC,        to murine HSCs, and not to human BM or UCB
human to immundeficient NOD/SCID mouse repopulating cell assay           HSC expansion, the application to human UCB

Hematology 2004                                                                                                        361
expansion could be very direct.                                  ment to ANC > 500 was seen on day 24–33, and plate-
      More recently, Zhu et al have discovered that the          let engraftment to > 50,000 at approximately day 90.
protein NF-Y is a normal transcriptional activator for                These results, while disappointing, highlight the
multiple Hox genes, as well as telomerase, Notch-1 and           importance of building clinical trial design based upon
LEF-1.16 When overexpressed in murine HSCs by as                 the most precise and appropriate preclinical data. The
little as twofold, NF-Ya increases HSC numbers by 20X            cytokine cocktails employed in the three published stud-
following transplantation. Application of TAT-NF-Y               ies included some but not all of the cytokines required
protein transduction to UCB HSC expansion thus of-               to achieve UCB HSC expansion in vitro, as measured
fers the attractive possibility of activating multiple HSC       by our best NOD/SRC assays: One trial used SCF and a
expansion pathways simultaneously. The absolute and              Tpo analogue, but not Flt3-l, and the other two used
relative efficiencies of these protein treatments for ex-        Flt3-l, but not Tpo or SCF, and neither included IL-6
panding human UCB HSCs will need to be evaluated                 or soluble IL-6 receptor. In addition, a lesser but possi-
directly in NOD/SCID mouse xenotransplants, and then             bly contributing variable was that in all three trials the
in appropriate Phase I clinical trials.                          ex vivo expanded cells were added 10–12 days later,
      Similar approaches using other soluble, revers-            thus mitigating any accelerating effect they might have
ible, biochemical stimulators of stem cell transcrip-            otherwise have achieved.
tion programs, including perhaps copper chelation,                    It must be pointed out that these trial design limita-
have also been proposed and are under preclinical                tions were known to the clinical investigators, but the
and clinical trial.                                              lack of availability of clinical grade cytokines, and pos-
                                                                 sibly also HSC selection devices, prevented the optimal
Clinical Experience with Ex Vivo                                 cocktails and culture conditions from being employed.
    Expanded Hematopoietic Cells                                 These difficulties would be surmounted if simpler ex-
Clinical trials of ex vivo culture UCB cells have begun          pansion conditions, requiring few added reagents and/
to be reported, but their interpretation should be tem-          or devices while still supporting HSC expansion, could
pered by the knowledge that the conditions of culture            be devised and employed.
used to date have not been shown to support USC am-
plification, only CFU expansion. At least three trials                What if UCB HSC Numbers Are Not Rate
evaluating the combination of unmanipulated with ex                       Limiting? Exploring the Null Hypothesis
vivo expanded UCB grafts have been published to date;                 Another possibility that must be considered is that in-
in all three studies expanded cells were added on day                 creasing UCB numbers will not, by itself, accelerate
10–12 to supplement unmanipulated cells. One study                    engraftment. Perhaps there is a qualitative “immatu-
involved the use of a static culture system in defined                rity” defect that prevents rapid engraftment. If so, then
media while the other two used a perfusion culture ex-                is there a way to circumvent this defect: providing a
pansion system. Although no toxicities were seen in                   maturation effect through some combination of in vivo
these studies, no significant acceleration of neutrophil              cytokine treatment, or perhaps ex vivo cell activation
or platelet engraftment was seen.17-19 Neutrophil engraft-            targeted at maturation? Viewed through this perspec-
                                                                                 tive, one can envision manipulations of the
                                                                                 non-HSC components of the UCB graft, in-
Table 4. Ex vivo approaches to progenitor and stem cell expansion.               cluding the mature hematopoietic cells, the
                                                                                 immune cells, antigen-presenting cells, and
                          Evidence for               Application to
Protocol                HSC Expansion                 Clinical Trials            mesenchymal stromal cell components of the
CD34  + selected cells,        +              Limited trials, without best       graft. Interactions between HSC and non-
high-dose cytokines                         combinations due to cytokine         HSCs could form the basis for potentially
                                           availability. No nadir reduction.     improved engraftment in mixed graft settings
                                                                                 as well.20 Each of these manipulations could
Low-dose cytokines,            ?            In trials, without best cytokine     be evaluated by comparing engraftment in
continuous perfusion                        combinations due to cytokine
                                           availability. No nadir reduction.
                                                                                 NOD/SCID mice, prior to clinical transplanta-
                                                                                 tion trials.
                                                                                      In summary, experience with hematopoi-
Direct transcriptional        +++                         Future
activation of HSC cycling:                                                       etic cell expansion to date supports the fun-
HOXB4, telomerase, NF-Y                                                          damental principle of experimental hematol-
                                                                                 ogy, that the best clinical results follow from
Abbreviations: HSC, hematopoietic stem cells                                     the most careful, comprehensive and creative

362                                                                                      American Society of Hematology
preclinical trials in the proper in vitro and in vivo mod-     T lymphocyte functions required for protection of the
els. For the future, this experience suggests concentrat-      HSCT recipient from infection. In particular, the unique
ing on ex vivo manipulations that can be shown to di-          aspects of UCBT, compared to either BMT or mobi-
rectly increase HSC numbers following NOD/SCID en-             lized peripheral blood stem cell transplantation
graftment and/or accelerate the pace of engraftment in         (PBSCT), will be described. Factors influencing the
these same models.                                             development of adequate immune function after HSCT
                                                               include those related to the donor progenitor cell popu-
  III. IMMUNE RECONSTITUTION AFTER UMBILICAL                   lations, pre-formed lymphocytes in the donor product,
        CORD BLOOD CELL TRANSPLANTATION                        the host microenvironment, pharmacological interven-
                                                               tions, and antigenic exposure. All clinically relevant
               Kenneth I. Weinberg, MD*                        stem cell sources include both progenitor cells and pre-
                                                               formed lymphocytes (Figure 1). Prethymic progeni-
One of the events necessary for the success of hemato-         tors may include HSC, common lymphoid progenitors
poietic stem cell transplantation (HSCT), regardless of        (CLP), committed T progenitors (CTP), whose T lym-
the source of stem cells, is the development of a func-        phoid progeny depend on the host thymus for their de-
tional immune system from donor-derived cells. The             velopment.2
production of adequate granulocytes, platelets, and red
blood cells usually occurs rapidly after HSCT. In con-         Development of Thymic Progenitors
trast, the ability to produce lymphocytes, especially T        Upon entry to the thymus, progenitor cells undergo
lymphocytes, is delayed. As a result, serious infection        expansion; differentiation events including V(D)J re-
in the first 1–2 years after HSCT occurs in about 50%          combination to generate functionally rearranged TCR
of uncomplicated transplants from histocompatible sib-         genes; and both positive and negative selection events
ling donors, and up to 80%–90% of recipients of                that cull potential T lymphocytes that would either be
matched unrelated donor (MUD) marrow transplants               unresponsive to antigenic peptides presented by self-
or histocompatible recipients who developed GVHD.1             major histocompatibility complex (MHC) antigens or
Infections attributable to poor lymphocyte function in-        overly responsive to either MHC or self antigens. The
clude viral pathogens such as the herpes group (cytome-        ultimate fate of negatively selected cells is apoptosis.
galovirus [CMV], herpes simplex virus [HSV], vari-             Since most thymocytes are fated to die, maintenance of
cella zoster virus [VZV], human herpes viruses [HHV]
6 and 8), respiratory viruses (respiratory syncytial vi-
rus [RSV], parainfluenza viruses), adenovirus, and en-
teroviruses (Coxsackie and ECHO viruses), as well as
EBV-associated lymphoproliferative disorders. Other
infections that may result from inadequate cellular im-
munity include fungal infections such as Pneumocystis
carinii, Candida, and Cryptococcus. Increased suscep-
tibility to Aspergillus infection may reflect defects in
innate immunity. Susceptibility to some bacterial in-
fections, notably those caused by encapsulated organ-
isms (Staphylococcus pneumoniae, Haemophilus
influenzae), is determined by the inability to produce
antipolysaccharide antibodies.
     The emphasis of this review will be the relation-
ship between stem cell source and the development of

                                                               Figure 1. Two pathways for generation of T lymphocytes
* Children’s Hospital of Los Angeles, 4650 Sunset Boulevard,   after hematopoietic stem cell transplantation (HSCT).
Box 62, Los Angeles CA 90027                                   Shown at top is the adoptive transfer of post-thymic T cells
                                                               which then undergo expansion in the host, resulting in a limited
Acknowledgments: Supported by NIH grants R01 HL54729,          repertoire of mature T lymphocytes that can rapidly confer
HL70005, AI50765; P50 HL54850; M01 RR00043. The                antigen-specific responses, including graft-versus-host
                                                               disease (GVHD). Below is shown the pathway for differentia-
thoughtful input of Gay Crooks, Bruce Blazar, Robertson
                                                               tion of pre-thymic progenitors by migration into the recipient
Parkman, Nelson Chao, Wes Brown, Hisham Abdel-Azim and         thymus, resulting in the slow generation of a diverse repertoire
Dan Douek is greatly appreciated.                              that is host-tolerant.

Hematology 2004                                                                                                           363
thymic output depends on continual expansion and dif-         cell development. For example, in patients with severe
ferentiation of the immature prothymocytes, which lack        combined immune deficiency (SCID) receiving CD34+
expression of the TCR invariant CD3 complex, and the          haploidentical HSCT, the appearance of functional
co-receptors CD4 and CD8 (“triple negative,” TN).             mature T lymphocytes in the peripheral blood is 90–
     The major signals for expansion of immature              120 days after transplant, a time period similar to that
prothymocytes are two cytokines, interleukin-7 (IL-7)         required for normal prenatal immune ontogeny.
and c-kit ligand (KL), which are produced by thymic
epithelial cells (TEC). Murine studies have demonstrated      Post-Thymic T Lymphocytes
that pre-HSCT radiotherapy and chemotherapy kill TEC,         In contrast to prethymic progenitors, post-thymic T cells
resulting in defective intrathymic IL-7 production.           have already undergone maturation and have antigenic
Early studies demonstrated that the thymic microenvi-         and functional specificities that were conferred by the
ronment is a target of GVHD, and more recent studies          donor’s thymus. Donor-derived post-thymic T cells ex-
have shown that TEC organization and function is ab-          press a fixed TCR and thereby are programmed to re-
normal in experimental GVHD models. In non-HSCT               spond to a set of antigenic determinants. Although adop-
studies, aging also results in TEC damage and decreased       tive transfer of mature T lymphocytes had long been
intrathymic IL-7 production. Thus, thymic function may        thought to be a relatively static process, recent studies
be adversely affected by several phenomena commonly           have indicated that mature T lymphocyte populations
associated with clinical HSCT-cytotoxic therapies given       are dynamic and regulated by interactions with other T
pre-HSCT, GVHD, and recipient aging. Clinical stud-           lymphocytes as well as the host microenvironment.
ies have demonstrated that decreased capacity for pro-        Mature T lymphocytes may undergo activation if they
duction of new T lymphocytes is associated with GVHD          encounter a stimulating antigen and the proper set of
and increasing recipient age.3,4 The effects of pre-trans-    co-stimulatory signals. Activation of T lymphocytes
plant conditioning on thymopoiesis has been more dif-         leads to acquisition of functional properties such as cy-
ficult to assess because of the complexity of the prob-       tolytic capacity or cytokine secretion. Activated T cells
lem of dose intensity as well as the effects of previous      may proliferate and mature into memory cells, undergo
therapy for malignancies. Any assessment of the effect        apoptosis (activation-induced cell death [AICD]), or
of stem cell source, e.g., UCB, on immune reconstitu-         be anergized. Expansion of activated T lymphocytes is
tion must take into account the function of the thymic        dependent on production of IL-2 by helper T lympho-
microenvironment.                                             cytes and expression of high affinity IL-2 receptors (IL-
     The biology of prethymic progenitors is also highly      2R) by the responsive helper and cytotoxic cells.
relevant to the outcome of HSCT. The lack of pre-                  Recent studies have demonstrated that naive T lym-
existing TCR rearrangements means that donor-derived          phocytes in a lymphopenic environment can undergo
prethymic progenitors are unlikely to cause GVHD, as          proliferation while maintaining a mainly naive pheno-
long as the recipient thymus maintains the ability to         type.5 Such homeostatic proliferation differs from acti-
select out undesirable T cells. However, the develop-         vation-induced proliferation in which naive T cells rec-
ment of an immune repertoire from prethymic progeni-          ognize foreign antigens and proliferate in response to
tors may be delayed by ontogenic factors such as com-         IL-2. In homeostatic proliferation, the stimuli recog-
mitment to the T lymphoid lineage and the time nor-           nized by the TCR are the same self antigens that nor-
mally required for development of large numbers of            mally induce positive selection in the thymus. Like
diverse T cells. Between 7 to 9 weeks of gestation, he-       thymopoiesis, homeostatic proliferation also requires
matopoietically derived cells enter the thymic anlage,        IL-7, although the source is probably extrathymic, not
which forms from mesenchyme and endoderm of the               TEC. Homeostatic proliferation has also been described
III-IV pharyngeal pouches. Alloreactive T cells are           for memory T lymphocytes with evidence that IL-15 is
present by week 14–15 and antigen-specific T lympho-          critical for the expansion of these cells.
cytes appear later in the second trimester. Recapitula-            The appearance of T lymphocytes after HSCT is
tion of fetal ontogeny provides a framework for under-        conventionally seen as evidence of de novo generation
standing the development of lymphoid cells after HSCT.        of T lymphocytes from progenitor cells. However, as
The generation of T lymphocytes post-HSCT requires            previously noted, transplantation of stem cell products
events similar to those observed in normal immune de-         that have been extensively depleted of mature T lym-
velopment, e.g., lymphoid commitment, thymic mi-              phocytes in order to prevent GVHD results in signifi-
gration and entry, and thymic differentiation. It is likely   cant delays in immune reconstitution. Therefore, it is
that the time period required for such development is         likely that much of what has been attributed to T cell
similar to the time normally required for prenatal T          generation in the first few months after transplant is the

364                                                                                American Society of Hematology
result of adoptive transfer of post-thymic cells. Such            ence may be the age of the HSC—a few months for
cells may undergo homeostatic proliferation in the                UCBC versus decades for adult HSC. There have been
lymphopenic environment of the host, or IL-2 driven               some studies of the aging process in hematopoietic pro-
proliferation if activated by exposure to alloantigens or         genitors, but there is little direct evidence that HSC
nominal antigens. Analyses of the length of time re-              from adult donors have decreased capacity to contrib-
quired for de novo generation of T lymphocytes after              ute to lymphocyte development compared to other pro-
HSCT are best performed in patients receiving highly              genitors.
purified progenitors are depleted of all post-thymic T
lymphocytes.                                                      The Placenta as Unique Microenvironment
                                                                      That Influences UCBT
UCBC HSC                                                          Besides intrinsic differences, HSC in UCBC have had a
HSC derived from UCBC differ from those of adult                  different set of microenvironmental exposures compared
marrow or PBSCT in several ways. The most notable                 to those of adult marrow or PBSC. All HSC sources
difference is quantitative—the cell dose in UCBT is               are influenced by the microenvironment from which
significantly less than the HSC dose in adult sources             they are derived. An example of differences between
such as PBSC or marrow. While low cell dose has been              sources are some of the observed changes in HSC cell
associated with failure of hematopoietic engraftment,             cycle status, gene expression and adhesive and invasive
HSC dose may also influence immune reconstitution.                properties induced by mobilization procedures used to
In a murine congenic transplant model, increasing the             generate PBSC, e.g., G-CSF.
dose of phenotypic HSC administered resulted in greater                The placenta is a complex organ that regulates ma-
thymic cellularity post-transplant, as well as greater            ternal-fetal interactions. Many cytokines that can influ-
numbers of mature T lymphocytes derived from the                  ence lymphohematopoietic development, e.g., G-CSF,
transplanted cells.6 Thus, one of the limiting factors for        c-kit ligand (stem cell factor [SCF]), GM-CSF, IL-15,
T lymphocyte reconstitution after HSCT, besides mi-               and others, are produced by the placenta. Production of
croenvironmental defects, may be the number of pro-               G-CSF by the placenta may be especially relevant to
genitor cells entering the thymus and contributing to             UCBT (Figure 2). G-CSF is produced both by the
thymopoiesis. Limited numbers of HSC in UCBC com-                 maternal decidua and the fetal chorionic villi8 and en-
pared to marrow or PBSC products may delay or de-                 ters the fetal circulation by a process that does not re-
crease thymic regeneration.                                       quire a functional G-CSF receptor. G-CSF from the
     There are also qualitative differences between               mother probably does not enter the fetal circulation as
UCBC and other HSC sources (Table 5). Comparisons                 administration of recombinant human G-CSF (rhG-
of CD34+ CD38– phenotypic HSC derived from UCBC                   CSF) to pregnant macaques did not result in detectable
versus adult marrow have shown that UCBC contain a                rhG-CSF in the fetuses. The function of placental G-
higher frequency of LTCICs and a higher percentage                CSF production is unknown; however, it may serve as
of cycling cells.7 UCBC-derived HSC also had greater              an immunoregulator that protects the mother and fetus
cloning efficiency and generative capacity. Thus, the             from each other’s allogeneic immune systems. G-CSF
HSC in UCBC may be both more primitive and more                   inhibits the ability of placental mononuclear cells to
capable of regenerating hematopoiesis in the recipient.           mediate cytotoxicity against allogeneic targets includ-
The contrasting biological properties may represent in-           ing choriocarcinoma cells. The production of G-CSF
trinsic or environmental differences in the UCBC and              and other cytokines capable of HSC mobilization, e.g.,
adult marrow HSC. The most likely intrinsic differ-               SCF, is likely to explain the high percentage of HSC in

Table 5. Properties of umbilical cord blood (UCB) cell populations relevant to immune responsiveness.

Hematopoietic Stem Cells             Dendritic Cells              T Lymphocytes
Decreased number                     Decreased immune activity    Decreased number
Increased cell cycling               Decreased co-stimulation     Naive phenotype
Increased generative capacity        Decreased IL-12 production   Resistance to activation
More immature?                                                    Decreased cytokine expression
                                                                  Skewing to Th2 responses (decreased IL-2, INF-γ production)

Abbreviations: IL, interleukin; INF, interferon

Hematology 2004                                                                                                          365
                                                                                     besides the logistical problem of trans-
                                                                                     planting the intrathymic ETP popula-
                                                                                     tion, it is unclear if such cells could
                                                                                     home and engraft after transplantation.
                                                                                     Murine CLP can be transplanted into
                                                                                     irradiated hosts and contribute to post-
                                                                                     transplant lymphopoiesis. Co-transplan-
                                                                                     tation of CLP with HSC is able to more
                                                                                     rapidly restore immune function than
                                                                                     transplantation of HSC alone, and can
                                                                                     result in increased protection of mice
                                                                                     from experimental challenge with mu-
                                                                                     rine CMV.9 Although human cells with
                                                                                     CLP-like features can be found abun-
                                                                                     dantly in UCBC (personal communica-
                                                                                     tion, Gay Crooks), rigorous compari-
                                                                                     son of the numbers and functional prop-
                                                                                     erties of CLP in UCBC versus marrow
                                                                                     or PBSC still need to be performed.
Figure 2. Placental granulocyte colony-stimulating factor (G-CSF)
production affects umbilical cord blood cells (UCBC).
                                                                                      UCBC Post-Thymic T Cells
G-CSF produced in the placenta may affect both fetal HSC and dendritic cells.
Placental G-CSF may induce mobilization of the fetal HSC, resulting in significant    Studies over the last 20 years have dem-
numbers of circulating HSC among UCBC, which declines rapidly after delivery.         onstrated differences between neonatal
G-CSF may also suppress the immunological potency of fetal dendritic cells by         T lymphocytes and those of adults
interfering with IL-12 production and co-stimulatory activity. As a result, the
immune responsiveness of fetal T lymphocytes is attenuated and skewed away            (Table 5). The major notable difference
from Th1 responses.                                                                   between the T lymphocytes in UCBC
                                                                                      and those derived from adult marrow
                                                                                      or mobilized PBSC is the maturational
UCBC and their rapid disappearance from the neonate’s               status. Because the fetus is exposed to few foreign anti-
circulation after birth. In contrast to mobilization in-            gens, the T lymphocytes in UCBC are almost exclu-
duced by a short period of monotherapy with G-CSF                   sively naive. Naive T lymphocytes express a phenotype
before PBSCT, the HSC in UCBC have been exposed                     that is identifiable as CD45RA+ CD45R0– and CD62L+.
continually to a complex mixture of cytokines that likely           As individuals age, there is an increase in the frequency
affect their behavior, e.g., induce a high rate of cell             of T cells that have differentiated into a memory phe-
cycling,8 or alter the homing and invasive properties               notype, CD45RA– CD45R0+ and CD62L–/low as a result
needed for entry of progenitors into the thymus.                    of antigenic exposure. Memory cells are more readily
                                                                    activated by antigen than naive T cells and may be able
UCBC and Lymphoid Progenitors                                       to respond to weaker co-stimulatory signals through
Little is known about the numbers and functional prop-              the B7-CD28 pathway. The predominant naive pheno-
erties of circulating lymphoid progenitors present in               type of T lymphocytes in UCBC may contribute to the
UCBC compared to other sources. Commitment to lym-                  reduced alloreactivity observed after UCBT.
phoid differentiation by donor HSC could be a rate-                      Investigations of neonatal T cell function have been
limiting step in the ability to repopulate the recipient            driven by interest in the increased susceptibility to se-
immune system after HSCT. At present, there is some                 vere viral infections in neonates. For example, new-
controversy regarding the nature of intermediate lym-               borns infected with HSV have higher rates of dissemi-
phoid progenitors. The murine common lymphoid pro-                  nation (sepsis, meningoencephalitis) than adults with
genitor (CLP) has been described as a phenotypic popu-              primary infection, and increased severity of some viral
lation in the marrow that can contribute to T, NK, and              infections such as enteroviruses is observed until at least
B lymphopoiesis, but it is not clear if CLP generate all            several months of age. Investigations of neonatal helper
thymocytes. There is some evidence that early thymic                T lymphocytes have demonstrated decreased produc-
progenitors (ETP) in the thymus can arise in a CLP-                 tion of Th1 cytokines needed for cytolytic and anti-
independent manner. At present, CLP are the only lym-               viral responses, compared to adult T lymphocytes.10
phoid progenitor that are relevant to transplantation;              Specifically, the activation of the interferon-γ (IFN-γ )

366                                                                                       American Society of Hematology
gene is markedly reduced in neonatal T lymphocytes,        functions of DC. Recent data on DC generation after
because of decreased induction of transcription factors,   HSCT suggests that G-CSF administration significantly
e.g., NF-AT, which are required for activation of the      decreases the ability to produce IL-12.11 The decrease
IFN-γ gene after TCR stimulation. A central question       in IL-12 production after post-HSCT G-CSF adminis-
is whether such differences are intrinsic to UCBC ver-     tration may be relevant to UCBT. As discussed above,
sus adult T lymphocytes. Interpretation of the differ-     cells in the fetal circulation are probably constantly
ences has been confused by comparison of signaling         exposed to placental G-CSF. Furthermore, many clini-
properties of UCBC T lymphocytes to unfractionated         cal UCBT protocols have used post-transplant G-CSF
adult T lymphocytes. Since adult T lymphocytes are         administration to accelerate the development of granu-
comprised of both naive and memory T cells, while          locytes. The routine administration of G-CSF, com-
UCBC are almost exclusively naive T cells, ascribed        bined with intrinsic defects in DC function, may de-
differences between them may simply reflect differ-        crease the generation of Th1 responses after UCBT.
ences between the stringent activation requirements of     The expected consequences of such an effect would be
naive helper T lymphocytes and the more easily acti-       decreased capacity to control opportunistic viral and
vated memory cells. Only comparisons between UCBC          fungal infections.
and purified naive T lymphocytes from adult blood can
be used to evaluate whether UCBC T cells have unique       Anticipated and Empiric Properties of UCBT
properties different from adult peripheral blood naive     Based on the progenitor and immunological properties
T lymphocytes. Regardless of the reason, T lympho-         of UCBC, certain predictions can be made regarding
cytes from UCBC appear to be less capable of mediat-       the expected immune reconstitution after UCBT. Com-
ing Th1 cytotoxic responses than those derived from        pared to the detailed information available about the
adult sources.                                             recovery of marrow function and the decreased risks of
     The decreased capacity for Th1 responses has both     severe GVHD, there are few systematic data on lym-
positive and negative implications for HSCT. The de-       phoid recovery, and even less on such properties as an-
creased absolute numbers and decreased responsiveness      tigenic repertoire and effector function. As discussed
of UCBC T lymphocytes contribute to the decreased          above, immune function after HSCT can be derived
alloreactivity and greater tolerance for MHC disparity     from either the adoptive transfer of preformed mature
observed in clinical UBCBT. However, the decreased         T lymphocytes in the stem cell product or from the
responsiveness probably means that there is also less      development in the host of donor-derived lymphocytes
adoptive immunity derived from UCBC T lymphocytes.         from the donor HSC or other prethymic populations,
There are fewer T cells overall, and the number of         e.g., CLP.2 The adoptively transferred T cells would be
memory cells capable of mediating recall responses to      expected to represent most of the lymphocytes seen in
antigen is extremely limited. Any adoptive immunity        the first few months after transplant, but are more likely
must be derived from the naive T lymphocytes present       to be both impermanent and narrow in repertoire. Adop-
in the UCBC product. As discussed above, naive T lym-      tively transferred T cells after UCBT are largely naive
phocytes are difficult to activate, rendering them less    cells which will be less responsive to specific antigen
capable of responses to pathogens. These properties of     than the memory cells in adult sources and probably
the mature T lymphocytes in UCBC would predict that        less able to mediate Th1 effector functions than mar-
antiviral and fungal immunity may be decreased after       row or PBSC. The HSC and pre-thymic progenitors
UCBT than after BMT or PBSCT.                              are expected to produce a long-lasting, broad immune
                                                           repertoire but would be slow to develop. The increased
UCBC Dendritic Cells                                       generative capacity of UCBC HSC might predict more
Besides intrinsic differences in T lymphocytes, there      rapid engraftment kinetics, but would be expected to
may be differences in antigen presenting cells (APC)       be offset by the decreased dose of transplanted HSC in
derived from UCBC versus adult sources (Table 5).          the average UCBT. Analyses of UCBT recipients have
Analyses of dendritic cells (DC) in UCBC have shown        demonstrated slow de novo generation of T lympho-
that both so-called myeloid and plasmacytoid DC are        cytes in the first year after transplant; however, by 2
present, although there may be a decreased proportion      years post-UCBT recipients have evidence of both a
of myeloid DC compared to adult peripheral blood.          broad repertoire and thymic function.3,12,13 It is unclear
The myeloid dendritic cells are thought to promote Th1     if there are differences in the kinetics of post-transplant
responses by producing IL-12. Studies of the DC de-        lymphopoiesis in UCBT versus BMT. More rapid re-
rived from UCBC have indicated that there may be in-       covery in children versus adults and nonmyeloablative
trinsic defects in production of IL-12 as well as other    than myeloablative conditioning suggests that damage

Hematology 2004                                                                                                  367
to the thymic microenvironment as a result of aging or       virus infections after UCBT versus BMT or PBSCT
cytotoxic therapies may be important determinants of         need to analyze at risk populations, not the total num-
post-HSCT immune reconstitution.12                           ber of recipients. For several infections, notably CMV
     Although several studies have demonstrated the im-      and HHV-6, there is evidence that at-risk recipients may
portance of UCBC CD34+ cell dose for likelihood of           have greater rate of progression to symptomatic infec-
hematopoietic engraftment and engraftment kinetics,          tion, with decreased evidence of viral control, although
analyses of the engraftment kinetics of lymphoid popu-       this is not a universal finding.17-19 In the comparison of
lations are more limited. In a study from Case Western       UCBT with MUD transplants at Case Western Univer-
University, unrelated donor UCBT was compared to             sity, the overall risk of infection, including bacterial
the matched unrelated donor marrow transplantation.14        infection, correlated with the degree of lymphopenia.17
UCBT led to decreased rate of appearance of lymphoid         The risk of early bacterial infection in UCBT recipi-
populations in the first 3 months after transplantation.     ents may reflect differences in duration of neutropenia,
In the subsequent year, the absolute lymphocyte count        not just lymphocyte numbers or function.16
of the UCBT recipients exceeded that of the MUD re-               The available information about immune reconsti-
cipients. Indeed, the mean ALC in the UCBT group             tution after clinical UCBT is quite limited. In order to
was normal by day 200 post-transplant but remained           better understand both the kinetics and quality of im-
abnormally low in the MUD recipients during the en-          mune reconstitution after UCBT, analyses of specific
tire first year. Since the T lymphocytes seen in the first   lymphoid populations, immunological repertoire, mi-
100 days post-transplant are likely to be derived from       togen and antigen-driven proliferation and function, and
adoptively transferred mature T cells, the decreased         responses to test neo-antigens, e.g., keyhole limpet
lymphocyte count in the first 3 months after UCBT            hemocyanin, are needed. Ideally, such studies would be
would be expected as a result of lower amount of such        multi-institutional but use similar degrees of matching,
cells in UCBC products. The increased numbers of lym-        CD34+ cell dose, pretransplant conditioning, support-
phocytes observed later could reflect greater capacity       ive care and post-transplant immune suppression to con-
of the UCBC progenitors to produce lymphocytes de            trol for the variables of GVHD, conditioning related
novo. However, this is uncertain: an alternative expla-      microenvironmental damage and progenitor cell dose
nation could be that the increased GVHD observed in          that likely have an impact on immune reconstitution.
MUD recipients led to more severe lymphopenia than           At this time, the lack of direct comparisons of immune
that seen after UCBT.                                        reconstitution hinders the ability to make decisions re-
     The differences in UCBC T cells from adult sources      garding the optimal source of HSC for transplantation.
would be expected to translate into UCBT recipients          In a patient with an active viral infection, e.g., CMV or
having less GVHD but also less protection from viral         a history of Epstein-Barr virus lymphoproliferative dis-
infections such as CMV or EBV-LPD. Overall, the risk         ease (EBV-LPD), unrelated marrow from an immune
of infectious death in studies of UCBT may be higher         donor may be better than UCBT. However, potential
than that of transplant from other sources, but large,       HSCT recipients may have limited choices regarding
multi-institutional, randomized studies have not been        potential HSC sources, and frequently the ability to rap-
done, and there is a bias toward use of UCBT in sicker       idly perform transplant may trump any immunological
patients who cannot await a MUD BMT.15-19 In a non-          disadvantages of UCBT.
randomized study of children with ALL, the incidence
of viral infections was greater after UCBT than after        Strategies to Overcome the Problems
unrelated BMT.16 Complicating the analysis of actual             of Immune Reconstitution After UCBT
infectious risk for some infections like herpes viruses is   A paradox of UCBT is that some of the associated im-
the difference in potential source of virus between          munological problems are causally related to the prop-
UCBT and either BMT or PBSCT. CMV- or EBV-in-                erties that make UCBT attractive in the first place. For
fected B cells can be derived from either the donor or       example, the decreased alloreactivity that has been clini-
recipient in BMT or PBSCT. In UCBT, CMV- or EBV-             cally observed in UCBT reflects the quantitative and
infected cells would be expected to exclusively arise        functional abnormalities that make adoptive transfer of
from the recipient. Assuming that CMV-negative blood         antiviral immunity so unlikely immediately after UCBT.
products are administered to CMV-negative recipients,        Efforts to improve immune reconstitution after any form
then only CMV-positive recipients of UCBT are at sig-        of HSCT have been directed at two somewhat different
nificant risk of CMV disease, while CMV-negative BMT         questions—accelerating the rate of development of func-
or PBSCT recipients could be infected through their          tional immunity, especially that required to control cer-
seropositive donor. Therefore, comparisons of herpes         tain early infections, e.g., CMV; or altering the long-

368                                                                               American Society of Hematology
term qualitative and functional defects such as              not been reproducibly observed in larger animal mod-
hypogammaglobulinemia that contribute to morbidity           els and IL-7 has additional effects on mature T cells
and mortality over the first decade after transplant. The    that could limit its usefulness in allogeneic HSCT. Be-
first goal has been more pressing because of the high        sides promoting the development of thymocytes, IL-7
impact of opportunistic infection on transplant-related      treatment also increases the expansion of mature T lym-
mortality. Strategies in BMT or PBSCT designed to            phocytes by promoting their proliferation and survival.
increase the adoptive transfer of antigen-specific T cells   These effects have led to increased GVHD in some pre-
directed against important pathogens such as CMV or          clinical experiments but have not been observed by oth-
EBV have used co-transplantation of cloned T cells that      ers. Experimental usage of IL-7 in clinical allogeneic
have been expanded in vitro. The application of such         transplants should probably be initially restricted to T
strategies to UCBT would require either cloned cells         cell–depleted grafts, not UCBT, until more is known
that are derived from the host or a third party, or tech-    about both efficacy and promotion of GVHD.
niques for in vitro priming of the naive T cells in the           Another strategy to address the problem of mi-
UCBC graft. Nonspecific activation of the T cells in         croenvironmental damage has been to decrease the tox-
the UCBC product would likely increase the risk of           icity of the conditioning regimen. Besides decreasing
GVHD.                                                        the intensity of conditioning, another approach is the
     Besides the manipulation of the preformed T cells       administration of a biological response modifier, re-
in a graft, other strategies that have been proposed for     combinant keratinocyte growth factor (KGF). KGF is
accelerating immune reconstitution have been aimed at        a mesenchymally derived member of the fibroblastic
increasing the rate at which prethymic cells contribute      growth factor family which interacts with an epithe-
to the mature T lymphocyte compartment, either by            lial-specific receptor. Administration of KGF in both
increasing the number of transplanted lymphoid pro-          preclinical experimental models and clinical trials has
genitors or by accelerating their development in vivo.       demonstrated decreased regimen-related toxicities such
One potential strategy that has been demonstrated in         as oral mucositis, but also less GVHD. KGF protects
murine models is the co-transplantation of committed         TEC in murine transplant and aging models, thereby
progenitors, such as CLP.9 Administration of 3000 CLP        increasing thymopoietic capacity and improving both
in addition to 500 HSC in lethally irradiated mice pro-      the immediate rate of lymphocyte recovery and me-
vided significant protection from experimental infec-        dium-term (3–4 months) immune reconstitution. The
tion with murine CMV, compared to HSC alone. Im-             clinical relevance of protection of murine thymopoiesis
pressively, small numbers of CLP provide levels of pro-      by KGF needs to be determined.
tection from MCMV that are comparable to that of mil-
lions of post-thymic cells. The technical difficulty for     Summary
UCBT would be obtaining sufficient CLP to affect re-         Immune reconstitution after UCBT is promoted by the
constitution, e.g., via ex vivo expansion. The use of        increased generative capacity of UCBC HSC, and the
ligands for the Notch receptors that regulate both pro-      decreased GVHD, but is hampered by fewer HSC, de-
liferation and commitment to lymphoid lineages might         creased adoptively transferred antigen-specific T cells,
be useful for in vitro generation of prethymic progeni-      and functional defects in the T cells. Clinical studies to
tors that could be used to augment those present in the      understand the relative importance of each of these bio-
UCBC product.20 Based on the data of Chen et al, expan-      logical properties to outcome are needed. At this time,
sion techniques to increase the number of transplanted       it is difficult to use relative differences in immune re-
HSC might be expected to increase thymic output.8            constitution between HSC sources as a basis for choos-
     Other strategies have focused on ways to increase       ing the mode of HSCT, although BMT might be pref-
the in vivo development of T lymphocytes in the re-          erential to UCBT for a recipient with an active viral
cipient after transplantation. These studies have been       infection. Some but not all strategies aimed at improv-
driven by the hypothesis that the thymic microenviron-       ing immune reconstitution after HSCT in general may
ment of HSCT recipients lacks sufficient capacity to         be relevant to addressing the immunological and infec-
efficiently produce new T lymphocytes from prethymic         tious problems observed after UCBT.
progenitors. Our group has emphasized that defects in
TEC numbers or function caused by aging, cytotoxic
therapies, or GVHD lead to a relative lack of intrathymic
IL-7 needed for thymopoiesis. Systemic administration
of IL-7 in murine models can dramatically improve
thymic output after transplantation, but the effects have

Hematology 2004                                                                                                   369
                          REFERENCES                                   18. Barker JN, Weisdorf DJ, DeFor TE, et al. Rapid and complete
                                                                           donor chimerism in adult recipients of unrelated donor
                                                                           umbilical cord blood transplantation after reduced intensity
I. Cord Blood Transplants: How Close Are We to                             conditioning. Blood. 2003;102:1915-1919.
    Using This in Adults?                                              19. McSweeney PA, Bearman SI, Jones RB, et al.
1. Ende M, Ende N. Hematopoietic transplantation by means of               Nonmyeloablative hematopoietic cell transplant using cord
    fetal (cord) blood. Virginia Med Monthly. 1972;99:276-280.             blood [abstract]. Blood 2001;98:666a.
2. Gluckman E, Broxmeyer HA, Auerbach AD, et al. Hemato-               20. Chao NJ, Liu CX, Rooney B, et al. Nonmyeloablative regimen
    poietic reconstitution in a patient with Fanconi’s anemia by           preserves ‘niches’ allowing for peripheral expansion of donor
    means of umbilical-cord blood from an HLA-identical                    T cells. Biol Blood Marrow Transplant. 2002;8:249-256.
    sibling. N Engl J Med. 1989;321:1174-1178.                         21. Rocha V, Labopin M, Frasoni F, et al. Results of unrelated
3. Kernan NA, Bartsch G, Ash RC, et al. Analysis of 462                    cord blood versus unrelated bone marrow transplant in adults
    transplantations from unrelated donors facilitated by the              with acute leukemia. Blood. 2002;100:148.
    National Marrow Donor Program. N Engl J Med.                       22. Laughlin M, Rubinstein P, Stevens C, et al. Comparison of
    1993;328:593-602.                                                      unrelated cord blood and unrelated bone marrow transplants
4. Barker JN, Krepski TP, DeFor TE, et al. Searching for                   for leukemia in adults: a collaborative study of the Interna-
    unrelated donor hematopoietic stem cells: availability and             tional Bone Marrow Transplant Registry and the New York
    speed of umbilical cord blood versus bone marrow. Biol                 Blood Center. Blood. 2003;103:857.
    Blood Marrow Transplant. 2002;8:257-260.
5. Gluckman E, Rocha V, Chevret S. Results of unrelated                II. Expansion of Umbilical Cord Blood Cell
    umbilical cord blood transplant. Transfus Clin Biol.                   Progenitors and Stem Cells: Biology and
6. Rocha V, Wagner JE Jr, Sobocinkski KA, et al. Graft-versus-             Application to Clinical Transplants
    host disease in children who have received a cord blood or         1. Broxmeyer HE, Douglas GW, Hangoc G, et al. Human
    bone marrow transplant from an HLA-identical sibling. N                umbilical cord blood as a potential source of transplantable
    Engl J Med. 2000;342:1846-1854.                                        hematopoietic stem/progenitor cells. Proc Nat Acad Sci U S A.
7. Laughlin MJ, Barker J, Bambach B, et al. Hematopoietic                  1989;86:3828-3832.
    engraftment and survival in adult recipients of umbilical-cord     2. Carow CE, Hangoc G, Cooper SH, Williams DE, Broxmeyer
    blood from unrelated donors. N Engl J Med. 2001;344:1815-              HE. Mast cell growth factor (c-kit ligand) supports the growth
    1822.                                                                  of human multipotential progenitor cells with a high replating
8. Sanz GF, Saavedra S, Planelles D, et al. Standardized,                  potential. Blood. 1991;78:2216-2221.
    unrelated donor cord blood transplantation in adults with          3. Holyoake TL, Nicolini FE, Eaves CJ. Functional differences
    hematologic malignancies. Blood. 2001;98:2332-2338.                    between transplantable human hematopoietic stem cells from
9. Rocha V, Arcese W, Sanz G, et al. Prognostic factors of                 fetal liver, cord blood and adult marrow. Exp Hem.
    outcome after unrelated cord blood transplant (UCBT) in                1999;27:1418-1427.
    adults with haematologic malignancies [abstract]. Blood.           4. Rocha, V, Wagner JE, Sobocinski K, et al. Comparison of
    2000;96:587a.                                                          graft-vs-host disease in children transplanted with HLA
10. Goldberg SL, Chedid S, Jennis AA, Preti RA. Unrelated cord             identical sibling umbilical cord blood vs HLA identical sibling
    blood transplantation in adults: a single institution experience       bone marrow transplant. N Engl J Med. 2000;342:1847-
    [abstract]. Blood. 2000;96:208a.                                       1854.
11. Cornetta K, Laughlin M, Carter S, et al. Umbilical cord blood      5. Uchida N, Aguila HL, Fleming WH, Jerabek L, Weissman IL.
    transplantation in adults: results of a prospective, multi-            Rapid and sustained hematopoietic recovery in lethally
    institutional, NHBLI sponsored trial [abstract]. Blood.                irradiated mice transplanted with purified Thy-1.1lo Lin-Sca-
    2002;100:42a.                                                          1+ hematopoietic stem cells. Blood. 1994;83:3758-3779.
12. Ooi J, Iseki T, Takahashi A, et al. Unrelated cord blood           6. BitMansour A, Burns SM, Traver D, et al. Myeloid progeni-
    transplantation for adults patients with de novo acute myeloid         tors protect against invasive aspergillosis and Pseudomonas
    leukemia. Blood. 2004:103:489-491.                                     aeruginosa infection following hematopoietic stem cell
13. Ooi J, Iseki T, Takahashi S, et al. Unrelated cord blood               transplantation. Blood. 2002;100(13):4660-4667.
    transplantation for adult patients with advanced                   7. Dorrell C, Gan OI, Pereira DS, Hawley RG, Dick JE.
    myelodysplastic syndrome. Blood. 2003;101:4711-4713.                   Expansion of human cord blood CD34(+)CD38(-) cells in ex
14. Long GD, Laughlin M, Madan B, et al. Unrelated cord blood              vivo culture during retroviral transduction without a
    transplantation in adult patients. Biol Blood Marrow                   corresponding increase in SCID repopulating cell (SRC)
    Transplant. 2003:9:772-80.                                             frequency: dissociation of SRC phenotype and function.
15. Rubinstein P, Carrier C, Carpenter C, et al. Graft selection in        Blood. 2000;95:102-110.
    unrelated placental/umbilical cord blood (PCB) transplanta-        8. Larochelle A. Vormoor J. Hanenberg H. et al. Identification
    tion: influence and weight of HLA match and cell dose on               of primitive human hematopoietic cells capable of repopulat-
    engraftment and survival [abstract]. Blood. 2000;96:588a.              ing NOD/SCID mouse bone marrow: implications for gene
16. Rizzieri DA, Long GD, Vredenburgh JJ et al. Successful                 therapy. Nat Med. 1996;2:1329-1337.
    allogeneic engraftment of mismatched unrelated cord blood          9. Moore MA. Hoskins I. Ex vivo expansion of cord blood-
    following a nonmyeloablative preparative regimen. Blood.               derived stem cells and progenitors. Blood Cells. 1994;20:468-
    2001;98:3486-3488.                                                     479.
17. Chao NJ, Koh L-P, Long GD, et al. Adult recipients of              10. Van Zant G, Drubachevsky I, Rummel S, Koller MF, Emerson
    umbilical cord blood transplantation following non-                    SG. Expansion of hematopoietic progenitor cells from
    myeloablative preparative regimen. Biol Blood Marrow                   umbilical cord blood via continuous perfusion culture. Blood
    Transplant. 2004. In press.                                            Cells. 1994;20:482-491.

370                                                                                             American Society of Hematology
11. Lewis ID, Du J, Almeida-Porada J, Zanjani ED, Verfaillie CM.     6. Chen BJ, Cui X, Sempowski GD, Domen J, Chao NJ.
    Long-term repopulating cord blood stem cells are preserved           Hematopoietic stem cell dose correlates with the speed of
    after ex-vivo culture in a non-contact system. Blood.                immune reconstitution after stem cell transplantation. Blood.
    2001:3441-3449.                                                      2004;103:4344-4352.
12. Glimm H, Tang P, Clark-Lewis I, von Kalle C, Eaves C. Ex         7. Hao QL, Shah AJ, Thiemann FT, Smogorzewska EM, Crooks
    vivo treatment of proliferating human cord blood stem cells          GM. A functional comparison of CD34+ CD38– cells in cord
    with stroma-derived factor-1 enhances their ability to engraft       blood and bone marrow. Blood. 1995;86:3745-3753.
    NOD/SCID mice. Blood. 2002;99:3454-3457.                         8. McCracken S, Layton JE, Shorter SC, Starkey PM, Barlow
13. Zandstra PW, Conneally E, Piret JM, Eaves CJ. Ontogeny-              DH, Mardon HJ. Expression of granulocyte-colony stimulat-
    associated changes in the cytokine responses of primitive            ing factor and its receptor is regulated during the develop-
    human haemopoietic cells. Br J Haematol. 1998;101:770-778.           ment of the human placenta. J Endocrinol. 1996;149:249-
14. Antonchuk J, Sauvageau G, Humphries RK. HOXB4-induced                258.
    expansion of adult hematopoietic stem cells ex vivo. Cell.       9. Arber C, BitMansour A, Sparer TE, et al. Common lymphoid
    2002;109:39-45.                                                      progenitors rapidly engraft and protect against cytomegalovi-
15. Krosl J, Austin P, Beslu N, Kroon E, Humphries RK,                   rus infection after hematopoietic stem cell transplantation.
    Sauvageau G. In vitro expansion of hematopoietic stem cells          Blood. 2003;102:421-428.
    by recombinant TAT-HOXB4 protein. Nat Med. 2003;9:1428-          10. Lewis DB, Yu CC, Meyer J, English BK, Kahn SJ, Wilson CB.
    1432.                                                                Cellular and molecular mechanisms for reduced interleukin 4
16. Zhu J, Gianolla D, Zhang Y, Rivera A, Emerson SG. NF-                and interferon-gamma production by neonatal T cells. J Clin
    Ya,b,c interacts with USF1/2 to activate the HOXB4 promoter          Invest. 1991;87:194-202.
    in human hematopoietic cells and repress granulopoiesis.         11. Fagnoni FF, Oliviero B, Giorgiani G, et al. Reconstitution
    Blood. 2003;102:2420-2427.                                           dynamics of plasmacytoid and myeloid dendritic cell
17. Shpall EJ, Quinones R, Giller R, et al. Transplantation of ex        precursors after allogeneic myeloablative hematopoietic stem
    vivo expanded cord blood. Biol Blood Marrow Transplant.              cell transplantation. Blood. 2004;104:281-289.
    2002;8:368-376.                                                  12. Klein AK, Patel DD, Gooding ME, et al. T-cell recovery in
18. Pecora A, Stiff P, Jennis A, et al. Prompt and durable               adults and children following umbilical cord blood transplan-
    engraftment in two older adult patients with high risk chronic       tation. Biol Blood Marrow Transplant. 2001;7:454-466.
    myelogenous leukemia (CML) using ex vivo expanded and            13. Talvensaari K, Clave E, Douay C, et al. A broad T-cell
    unmanipulated unrelated umbilical cord blood. Bone Marrow            repertoire and an efficient thymic function indicate a
    Transplant. 2000;25:797-799.                                         favorable long-term immune reconstitution after cord blood
19. Jaroscak J, Goltry K, Smith A, et al. Augmentation of                stem cell transplantation. Blood. 2002;99:1458-1464.
    umbilical cord blood (UCB) transplantation with ex vivo-         14. Hamza NS, Lisgaris M, Yadavalli G, et al. Kinetics of myeloid
    expanded UCB cells: results of a phase 1 trial using the             and lymphocyte recovery and infectious complications after
    AastromReplicell System. Blood. 2003;101:5061-5067.                  unrelated umbilical cord blood versus HLA-matched
20. Barker JN, Weisdorf DJ, DeFor TE, Blazar BR, Miller JS,              unrelated donor allogeneic transplantation in adults. Br J
    Wagner JE. Rapid and complete donor chimerism in adult               Haematol. 2004;124:488-498.
    recipients of unrelated donor umbilical cord blood transplan-    15. Rubinstein P, Carrier C, Scaradavou A, et al. Outcomes among
    tation after reduced-intensity conditioning. Blood.                  562 recipients of placental-blood transplants from unrelated
    2003;102:1915-1919.                                                  donors. N Engl J Med. 1998;339:1565-1577.
                                                                     16. Rocha V, Cornish J, Sievers EL, et al. Comparison of
III. Immune Reconstitution After Umbilical Cord                          outcomes of unrelated bone marrow and umbilical cord blood
   Blood Cell Transplantation                                            transplants in children with acute leukemia. Blood.
1. Ochs L, Shu XO, Miller J, et al. Late infections after
                                                                     17. Wagner JE, Barker JN, DeFor TE, et al. Transplantation of
   allogeneic bone marrow transplantations: comparison of
                                                                         unrelated donor umbilical cord blood in 102 patients with
   incidence in related and unrelated donor transplant recipients.
                                                                         malignant and nonmalignant diseases: influence of CD34 cell
   Blood. 1995;86:13979-13986.
                                                                         dose and HLA disparity on treatment-related mortality and
2. Mackall CL, Gress RE. Pathways of T-cell regeneration in
                                                                         survival. Blood. 2002;100:1611-1618.
   mice and humans: implications for bone marrow transplanta-
                                                                     18. Tomonari A, Iseki T, Ooi J, et al. Cytomegalovirus infection
   tion and immunotherapy. Immunol Rev. 1997;157:61-72.
                                                                         following unrelated cord blood transplantation for adult
3. Weinberg K, Blazar BR, Wagner JE, et al. Factors affecting
                                                                         patients: a single institute experience in Japan. Br J Haematol.
   thymic function after allogeneic hematopoietic stem cell
   transplantation. Blood. 2001;97:1458-1466.
                                                                     19. Sashihara J, Tanaka-Taya K, Tanaka S, et al. High incidence
4. Storek J, Joseph A, Dawson MA, Douek DC, Storer B,
                                                                         of human herpesvirus 6 infection with a high viral load in
   Maloney DG. Factors influencing T-lymphopoiesis after
                                                                         cord blood stem cell transplant recipients. Blood.
   allogeneic hematopoietic cell transplantation. Transplantation.
                                                                     20. Allman D, Karnell FG, Punt JA, et al. Separation of Notch1
5. Schluns KS, Kieper WC, Jameson SC, Lefrancois L.
                                                                         promoted lineage commitment and expansion/transformation
   Interleukin-7 mediates the homeostasis of naive and memory
                                                                         in developing T cells. J Exp Med. 2001;194:99-106.
   CD8 T cells in vivo. Nature Immunol. 2000;1:426-432.

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