1 These studies were supported under Grants CA-23175.
CA-12800. and CA-15688 from the National Cancer Institute. and Grant
RR-00865 from the U.S. Public Health Service.
2 Robert Peter Gale is a Scholar of the Leukemia Society of America.
Departments of Medicine, Microbiology & Immunology and the Bone
Marrow Transplant Unit, UCLA School of Medicine. the Center for
the Health Sciences. Los Angeles, Calif. 90024. USA
Acute leukemia is a neoplastic disease characterized by the abnormal
proliferation and accum illation of immature hematopoietic cells.
Progress in our understanding of this disease is reviewed in this
volume. Significant recent advances in the therapy of acute leukemia
include high remission rates in both acute lymphoblastic (ALL) and
acute myelogenous leukemia (AML), the prevention of central nervous
system leukemia in ALL, and development of moderately effective
remission maintenance programs, particularly in ALL. Despite this
progress, approximately 50 per cent of patients with ALL and over
95 per cent of those with AML will eventually die of resistant leukemia.
Recent studies at our institution and others have clearly demonstrated
the feasibility of transplantation of normal hematopoietic stem
cells in man. In view of this, and because of the disappointing
results of chemotherapy in patients with acute leukemia who relapse,
we studied the potential role of bone marrow transplantation in
resistant acute leukemia. In this chapter I will briefly review
some basic aspects of the biology and immunology of marrow transplantation
and discuss its applicability to leukemia.
Cell Biology and Immunology of Bone Marrow Transplantation
The hematopoietic system is derived from pluripotent stem cells.
These cells have several inherent characteristics relevant to marrow
transplantation including self-renewal potential, differentiative
capacity, and the presence of histocompatibility antigens (HLA)
on their surface. It is clearly possible to transplant hematopoietic
stem cells in man. Requirements for engraftment include histocompatibility
matching between donor and recipient, immunosuppression to prevent
graft rejection, and a critical dose of marrow cells. The latter
may relate to the clinical setting under which transplantation is
performed rather than an inherent characteristic of stem cell(s)
since a single cell may be capable of repopulating a congenitally
anemic non-irradiated mouse under appropriate conditions. The human
major histocompatibility complex (MHC), referred to as HLA, has
been assigned to chromosome 6. The HLA locus has been further subdivided
into the HLA-A, B, C, and D subloci. The first three are commonly
defined by serologic technics while the HLA-D region is conventionally
studied in the mixed lymphocyte culture (MLC) test (for review see
[ 1 ]). Successful marrow transplants in man have been restricted
to HLAidentical siblings with few exceptions. While HLA is of prime
importance in determining graft outcome, other histocompatibility
systems are undoubtedly involved. Little is known regarding these
non-HLA systems and no attempt has been made to match for non-HLA
antigens in clinical transplantation. This factor probably accounts
for the high incidence of graft rejection and graft-versus-host
disease (GVHD). There is a high degree of polymorphism in the HLA
system. Since these antigens are inherited in a Mendelian fashion
as codominant alleles, there is a reasonable possibility (25 per
cent) of finding an HLA-identical donor within a family. In the
general population the probability is in the range of one in 10.000.
Because of this, most transplants have been performed between HLA-identical
siblings. Despite profound hematopoietic suppression, patients with
aplasia and acute leukemia are capable of rejecting allogeneic grafts.
Immunosuppression is therefore necessary to achieve sustained marrow
engraftment. Pretransplant immunosuppression, referred to as conditioning.
has utilized chemotherapy and radiation either singly or in combination.
Since doses used in these regimens are supralethal, rescue with
normal marrow is essen tial for survival. The transplant procedure
is relatively simple. Approximately one liter of bone marrow is
removed from the donor by aspiration from the posterior iliac crests.
A single cell suspension is prepared and infused intravenously to
the recipient. The infused cells home to the marrow after a brief
delay in the lungs and spleen. The usual dose is 1-5 X 10 high 8
nucleated marrow cells per kg. In most instances discrete clusters
(colonies) of hematopoiesis are observed in the marrow with the
first 2 weeks following transplantation . These clusters are
usually either erythroid or granulocytic, but mixed populations
are occasionally observed. Peripheral white blood cells and platelets
begin to rise within 2-3 weeks following transplantation and may
return to normal levels by 1-2 months. Cytogenetic and gene marker
studies clearly indicate that red cells, granulocytes, lymphocytes,
platelets, monocytes, and hepatic and alveolar macrophages are ofdonor
origin [4,14,16]. Following successful engraftment, the recipient
is at risk to develop several immune-related problems including
graft rejection, graft-versushost disease (GVHD), post-transplant
immunodeficiency, interstitial pneumonitis, and infectious complications
(Table 1). Recurrent leukemia is an additional potential complication
in leukemic recipients. Graft rejection probably results from histoincompatibility
between donor and recipient and may be facilitated by immunization
of the recipient via blood transfusions. In some instances defects
in the marrow microenvironment may be responsible for graft failure
. Graft failure occurs in
Table I. Areas of investigation
Graft rejection Resistant
leukemia Graft-vs-host disease
Infectious disease complications
20-40 per cent of patients with aplastic anemia but is rare in
patients with leukemia. This may relate to either inherent differences
between the two diseases or to the more intensive conditioning used
in leukemics. Storb and coworkers have reported a correlation between
graft rejection and both recipient anti-donor immunity and marrow
dose , and we have reported a correlation with pre-transplant
lymphocytotoxins . Transfusions also contribute to graft rejection.
Graft-versus-host disease results from the introduction of immunocompetent
donor cells into the immunosuppressed recipient. Principle target
organs of GVHD include the lymphoid system, skin, liver, and gastrointestinal
tract . GVHD in man results from incomplete matching for nonHLA
histocompatibility antigens. The loss of normal immune regulatory
mechanisms and autoimmunity may also contribute. While GVHD initially
results from immune stimulation, the end result is immunodeficiency.
The incidence of GVHD following HLA-identical marrow transplantation
is 70 per cent, and over one-half of these cases are fatal. The
prevention and treatment of GVHD are problematic. Methotrexate is
routinely given prophylactically to modify GVHD, but this is not
completely effective. Attempts to prevent GVHD with antithymocyte
globulin (A TG) or to treat it with A TG , corticosteroids, and
other imm unosuppressive drugs have been largely unsuccessful. While
complete histocompatibility matching would theoretically prevent
GVHD, this approach would further limit the number of potential
candidates for bone marrow transplantation. The removal of immunocompetent
cells from the marrow inoculum prior to transplantation by either
physical or immunologic technics has appeal but has not been critically
evaluated in man. The complete prevention of GVHD is not necessarily
desirable since GVHD may have anti-leukemic effects. Allogeneic
marrow transplantation is followed by a period of immunodeficiency
lasting several months to 1-2 years [7,20]. The cause of the immunodeficiency
is multifactoral and includes abnormal or delayed lymphoid differentiation,
GVHD, and the effects of immunosuppressive drugs. Posttransplant
immunodeficiency is characterized by abnormalities of both T and
B lymphocyte function including decreased antibody synthesis, decreased
responsiveness to polyclonal mitogens, and inability to be sensitized
to dinitrochlorobenzene (DNCB). Reactivity to alloantigens and skin
graft rejection are normal. This immunodeficiency is correctable
with time. This suggest that either a small number of lymphoid precursors
are engrafted. or that their development is delayed. We have found
no evidence of suppressor cells or factors in these patients .
Approximately 60-70 per cent of marrow graft recipients develop
interstitial pneumonitis [ II ]. The incidence is higher in leukemic
patients than in aplastics. One-half of cases are related to cytomegalovirus
(CMV), 10 per cent to pneumocystis, and 10 per cent to other viruses.
No etiology is identified in the remaining cases. It is likely that
immunologic factors including immune stimulation, immunodeficiency,
and GVHD playa critical role in the development of interstitial
pneumonitis. Radiation and/or chemotherapy are probably not primary
factors but may compromise resistance. In CMV pneumonitis, it is
likely that both reactivation of latent endogenous infection and
exogenous infection are important factors. Attempts to prevent or
treat interstitial pneumonitis with antiviral chemotherapy (ara-A)
have been unsuccessful. Studies of CMV immune globulin, or plasma
and interferon, are currently underway at several centers. Bacterial
and fungal infections are an important complication of bone marrow
transplantation . These usually occur during the period of granulocytopenia
immediately following the transplant and their magnitude is related
to the intensity of the conditioning regimen. Most patients receive
oral non-absorbable antibiotics for gastrointestinal tract sterilization.
systemic antibiotics, and granulocyte transfusions. The value of
prophylactic granulocyte transfusions and laminar air flow environments
is controversial, but recent data suggest they may decrease the
incidence of infection without a substantial effect on survival
Current Results in Acute Leukemia
The survival of patients with resistant acute leukemia is poor
with median survival of less than 6 months in several large series.
Because of this, we and others have studied the potential role of
allogeneic bone marrow transplantation in patients with resistant
disease. Transplantation in acute leukemia is difficult. In addition
to the previously described immunobiologic problems, it is necessary
to permanently eradicate the leukemic clone(s). A variety of chemotherapy-radiation
therapy regimens have been developed to achieve this goal. Three
representative regimens are indicated in Fig. I and remission and
survival data in Figs. 2 and 3 [8,9, 13, 18]. Several important
points emerge from these studies: I. Leukemic relapse is common
despite the use of supralethal levels of drugs or radiation; 2.
The risk of relapse is high during the first 2 years but lower thereafter;
3. That with the possible exception of SCAR! (see legend Fig. I),
more intensive conditioning has not been associated with a lower
relapse rate; and 4. 15-20 per cent of patients with resistant disease
may become long-term disease-free survivors. While this survival
rate is not a satisfactory end result. it is probably superior to
chemotherapy alone. It is noteworthy that immunologic problems rather
than resistant leukemia are the major cause of death in some series.
These problems may ultimately prove more soluble than resistant
Major features of bone marrow transplantation in leukemia are reviewed
in Table 2. It should be emphasized that 98 per cent of relapses
occur in recipient cells so that progress is dependent upon the
development of more effective conditioning regimens. Potential approaches
to this problem are indicated in Table 3. Perhaps the most promising
are the development of more effective regimens and transplantation
in remission. Preliminary data from Thomas and coworkers has indicated
a low relapse rate in patients transplanted in remission. Finally.
the introduction of new myelosuppressive drugs or innovative uses
of radiation may improve the results of transplantation in acute
Table 2. Leukemic recurrence following bone marrow transplantation
Indefinite time at risk
Predominantly in recipient cells
Maintains genetic markers of original disease
Residual normal hematopoiesis of donor origin
Table 3. Approaches to decreasing the rate of leukemic
More effective chemoradiotherapy
Intensive chemoradiotherapy with optimal support facilities
Treat leukemic "sanctuaries'.
Transplant before "resistance" develops
Combination of approaches
Future research in this field must concentrate on two critical
1. More effective leukemia eradication and, 2. solutions to immunologic
problems including GVHD, immunodeficiency, and interstitial pneumonitis.
With regard to the first area, I have discussed the development
of more effective conditioning regimens and transplantation in remission.
GVHD is a difficult problem and it now seems clear that prevention
is critical. Based on animal data. the selective elimination of
immunocompetent cells from the graft by either physical or immunologic
technics seems a logical step. Immunodeficiency is probably best
approached by steps to facilitate the rate of lymphoid maturation.
These might conceivably involve transplants of thymic epitheli um
of the use of thymic hormones. Progress in the area of interstitial
pneumonitis will depend on an understanding of the etiologic and
pathogenic mechanisms involved. Trials of CMV immune plasma and
interferon are currently in progress. Development of more effective
antiviral chemotherapy such as phosphonoacetic acid for CMV infection
is clearly needed. Progress in GVHD and immunodeficiency may have
a beneficial effect on interstitial pneumonitis. Finally, the possibility
of lungshielding should be considered in non-leukemic patients.
A recent area of considerable interest is autotransplantation using
cryopreserved remission bone marrow (for review see [ 10]). Preliminary
studies have clearly indicated that cryopreserved marrow can reconstitute
a lethally radiated recipient but leukemic relapse has been a major
obstacle. Whether this relates to residual leukemia in the patient
or in the cryopreserved marrow is as yet uncertain. The concept
of autotransplantation is of considerable theoretical interest since
these patients would not be at risk to develop many of the immunologic
problems associated with allogeneic transplantation such as GVHD.
Autotransplantation could expand the applicability of marrow transplantation
since most patients with leukemia lack an HLAidentical sibling donor.
Leukemic relapse remains the major problem in autotransplantation,
and at temps to deplete clinically undetectable leukemic cells from
remission marrow using either physical or immunologic technics need
to be critically evaluated. A final consideration is the use of
HLA-matched unrelated donors for patients without HLA-identical
siblings. Recent advances in histocompatibility testing, particularly
HLA-D typing, make this a possibility. Opelz and coworkers have
recently reviewed theoretical consideration involved in the development
of donor pools for unrelated marrow transplantation .
Bone marrow transplantation is an experimental approach to the
treatment of patients with acute leukemia, aplastic anemia, and
other neoplastic and genetic diseases. To date, long-term disease-free
survival has been achieved in a small proportion of carefully selected
patients with resistant acute leukemia. While results are not optimal,
they are acceptable in late stage patients where there are no effective
alternates. Major problems in marrow transplantation for leukemia
include tumor resistance and a spectrum of immunologic complications
including GVHD, immunodeficiency, and interstitial pneumonitis.
Potential approaches to these problems have been suggested. Progress
in anyone area would have a substantial impact on improving survival
and extending the applicability of marrow transplantation to patients
at an earlier stage of their disease.
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