British journrrl of Hn~rtrntology. 1 9x4. 56. 4 3 1-44 3 Cytogenetic studies on recipients of allogeneic bone marrow using the sex chromosomes as markers of cellular origin SYLVIA D. LAWLER, MAKION c. BAKER, HELEN 11AKKIS AND G. R. MOKGENSTERN* Sortions of Hirrnrin Ccnctics rind MidiciniJ of the lnstitiite of Ciinc.fr Rrsrardi and the Royal Mrirsdm Hospitcrl, Siirrey SIJMMAKY. In 45 patients whose donor was of unlike sex, the sex chromosomes were used as markers of the cellular origin of niyeloid and lymphoid tissues after allogeneic bone marrow transplantation (HM'I'). Successful engraftment was characterized by the appearance of dividing donor cells in marrow within 2 weeks of grafting and in mitogen stimulated blood cultures by 3 weeks. Leukaemic relapse was identified in eight cases and was associated with difierent patterns of cellular origin of the myeloid and lymphoid tissues. At the time of relapse the marrow contained either a mixed population of normal donor and leukaemic recipient cells, or only recipient cells. Thus, in this series. leukaemic relapse was not found occurring in donor cells. The importance of defining the origin of cells in interphase as well as in metaphase was demonstrated. In all but one case. the dividing lymphoid population remained of donor origin during relapse. In the Leukaemia Ilnitt at the Royal Marsden Hospital, bone marrow transplantation (HMT) from family members is used for the treatment of leukaemia. In this paper we describe the use of the sex chromosomes as markers to identify recipient and donor cells after BMT. This approach has given us an overall view of the origin of the cell populations in marrow and blood after grafting and has enabled us to interpret unusual observations in relation to the tindings in the majority of cases. * Leukaemia Kesearch Fund Fellow. Members of the Lcukacmia IJnit: K. I,. I'owIcs. T. J. Mctilwain. I). N. I,awson. Fi. E. M. Kay, H. M. Clink. H. Jameson. S. 11. Lawler. A. 13arrctt and 1). Rorlcy. Correspondence: Professor Sylvia I). I,awlcr. Koyal Marsden t{ospital. Pulham Koad. I,ondon SW 3 hJJ. 431
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Cytogenetic studies on recipients of allogeneic bone marrow using the sex chromosomes as markers of cellular origin
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British journrrl of Hn~rtrntology. 1 9x4. 56. 4 3 1-44 3
Cytogenetic studies on recipients of allogeneic bone marrow using the sex chromosomes as markers of cellular origin
SYLVIA D. L A W L E R , M A K I O N c. B A K E R , H E L E N 1 1 A K K I S A N D
G. R . MOKGENSTERN* Sortions of Hirrnrin Ccnctics r i n d MidiciniJ of the lnstitiite of Ciinc.fr Rrsrardi and the Royal Mrirsdm Hospitcrl, Siirrey
SIJMMAKY. In 45 patients whose donor was of unlike sex, the sex chromosomes were used as markers of the cellular origin of niyeloid and lymphoid tissues after allogeneic bone marrow transplantation (HM'I'). Successful engraftment was characterized by the appearance of dividing donor cells in marrow within 2 weeks of grafting and in mitogen stimulated blood cultures by 3 weeks. Leukaemic relapse was identified in eight cases and was associated with difierent patterns of cellular origin of the myeloid and lymphoid tissues. At the time of relapse the marrow contained either a mixed population of normal donor and leukaemic recipient cells, or only recipient cells. Thus, in this series. leukaemic relapse was not found occurring in donor cells. The importance of defining the origin of cells in interphase as well as in metaphase was demonstrated. In all but one case. the dividing lymphoid population remained of donor origin during relapse.
In the Leukaemia Ilnitt a t the Royal Marsden Hospital, bone marrow transplantation (HMT) from family members is used for the treatment of leukaemia.
In this paper we describe the use of the sex chromosomes as markers to identify recipient and donor cells after BMT. This approach has given us an overall view of the origin of the cell populations in marrow and blood after grafting and has enabled us to interpret unusual observations in relation to the tindings in the majority of cases.
* Leukaemia Kesearch Fund Fellow. Members of the Lcukacmia IJnit: K. I , . I'owIcs. T. J . Mctilwain. I). N . I,awson. Fi. E. M. Kay, H. M.
Clink. H. Jameson. S. 11. Lawler. A. 13arrctt and 1). Rorlcy.
Correspondence: Professor Sylvia I) . I,awlcr. Koyal Marsden t{ospital. Pulham K o a d . I,ondon SW 3 hJJ.
431
4 32 Sylviii D. Lnwler at id
PATIENTS
Of the 138 patients with leukaemia who were treated by BMT between November 1977 and June 1982, samples of bone marrow and/or blood were available at some stage from 45 patients. 22 males and 2 3 females, who had received marrow from a donor of unlike sex. In 40 cases the donor was an HLA-identical sibling and in five cases the donor was a sibling, parent or child who was matched for only one haplotype with the recipient. Most of the patients were prepared for bone marrow transplantation with cyclophosphamide and total body irradiation (TBI) but a few patients were given melphalan in high dosage a s an alternative to TBI. All the blood products given after BMT were irradiated at 1 5 Gy.
MF,THODS AND MATEKIAIS
The type of leukaemia was described according to the FA13 classification (Bennett rt nl, 1976). Bone marrow chromosome preparations were made using a modification of the direct and 24 h culture method of Tjio & Whang ( 1 962) and the cell synchronization technique of Yunis ( 1 97h).
Peripheral blood cultures were set up either from the buffy-coat layer or whole blood in M L ~ O Y ' S medium 5A (Gibco) supplemented with 20-30(%, autologous or fetal calf serum (Gibco). and with the addition of penicillin ( 1 0 0 ILJ/ml) and streptomycin (100 pg/ml). The cells were cultured ( a ) without mitogen and (b) with the following mitogens: phytohaeni- agglutinin M (PHA) (Wellcome) 1 : 100. pokeweed mitogen (PWM) (Gibco) 1: 1 0 0 and lipopolysaccharide (1,PS) from 1;. rdi 0 1 11 R 4 (Sigma) 40 pg/ml. The culture times varied from 2 to 7 d in different cases.
The chromosomes were G-banded using trypsin and Giemsa (GTG) and karyotypes were described according to the standard international nomenclature (ISCN. 1978).
Y-body assessment was made following staining in 0.5';/0 quinacrine dihydrochloride (Atebrin: Gurr) and mounting in pH 6 . 8 phosphate buffer (Gurr).
K ES IJ LTS
Ilonor cells can be identified in bone marrow within 2 weeks of grafting but no dividing recipient cells were found until 9 weeks after grafting. Such recipient cells were identitied in seven of the 39 patients from whom marrow samples were obtained.
Cytogenetic studies of peripheral blood mononuclear cells were also made at various times after grafting in 3 1 patients. Donor cells could be identified in blood by 3 weeks after grafting and no recipient cells were found, whether or not mitogen was present, until 8 weeks after grafting. Kecipient cells were identified in eight cases.
The clinical and cytogenetic details of the patients who had recipient cells in marrow and/or blood a t any time after grafting are shown in Table I. Each patient has been assigned B number in this series but the Royal Marsden Hospital 13MT number is also given so that the patient can be identified in other publications.
Cytogenetic Studies in Allogmric BMT 43 3
(1) Occasional recipient cells (three cases)
tn Case 1 one normal recipient cell was found 14 weeks after grafting in an unstimulated blood culture, so the cell could have belonged to the myeloid or lymphoid series. A single normal host cell was found 9 weeks after grafting in the marrow of Case 2. Both these patients subsequently died of graft versus host disease with no post-mortem evidence of leukaemia. A single normal recipient cell was also found 18 weeks after grafting in an LPS stimulated blood culture in Case 3 , who was well and in complete remission 39 weeks later.
( 2 ) Presence of recipient cells revealed by LPS mid I’WM (one c(ise)
Case 4, the only patient in this series with a diagnosis of B-cell ALL, was also the only case in which a proportion (one-fifth) of normal host cells was found in blood cultures stimulated by PWM and LPS. On the other hand. in the PHA stimulated cultures, almost all metaphases were of normal donor karyotype. although about one in 2 0 cells showed gross chromosomal damage and consequently the sex chromosomes could not be identified with certainty. The absence of normal undamaged recipient cells in the presence of PHA. and their occurrence in the cultures stimulated by the other mitogens, suggests that they were B cells. Details of the cultures are shown in Table I. The patient remains well and in complete remission.
( 3 ) Recipient cells in leukaemic relapse (e ight iwses)
In Case 5 the two recipient metaphases found 10 weeks after grafting in a PHA-stimulated culture showed gross radiation type damage. At this time there was no clinical evidence of relapse. Five months later when the patient was in relapse, only recipient cells were identified in the marrow in interphase and metaphase. Although the sex of these cells that presumably belonged to the leukaemic clone was determined, they could not be fully analysed; their chromosome number was near diploid. No information is available about the karyotype of the bone marrow prior to BMT.
Case 6, like Case 5, initially showed successful engraftment as demonstrated by the presence of only donor metaphases in the marrow. However, 6 I weeks after grafting, three recipient cells, only one of which had a normal karyotype, were found in the blood. Five weeks later the absence of normal donor cells in blood cultures was confirmed. It was not until 72 weeks after grafting that relapse was diagnosed and found to be associated with the presence of recipient cells in the marrow and a total absence of donor cells in myeloid or lymphoid series. This patient had received marrow from a brother who was matched for only one haplotype. Seven of the eight recipient metaphases found in the marrow at relapse could not be analysed. but one was pseudodiploid. A marrow examined when the recipient was in relapse before BMT yielded no analysable mitoses.
In Case 7 the ratio of recipient to donor cells of 1 : 3 shown in Table I is not representative. In Table I aneuploid cells without a Y, as determined by fluorescent studies, in which only a single X chromosome could be identified, have not been included. Such cells which have lost a sex chromosome can be interpreted as belonging to the recipient, or to the donor. The details
Tabl
e I.
Clin
ical
and
cyt
ogen
etic
det
ails
of
sele
cted
pat
ient
s ~~
~~
No.
of
reci
pien
t ce
lls
Rec
ipie
nt
,'No. o
f do
nor c
ells
Wee
ks
Serie
s U
MT
Dia
gnos
is
Con
ditio
ning
G
vHII
af
ter
Peri
pher
al
No.
N
o.
Age
. Sex
an
dsta
ge
Don
or
regi
me
prop
hyla
xis
BMT
Mar
row
bl
ood
Com
men
ts a
nd o
utco
me
1 34
35
M
AML(M1) Si
ster
Cy
,'TBI
1s
t CR
2 52
46
M
AML
(M1)
Sis
ter
Cy/
TBI
1st C
R
3 1
30
14
M
AM
L (M
2) S
iste
r w
Tn
I 1s
t CR
4 1
33
7
M
B-A
LL
Sist
er
Cy1T
BI
1st C
R
5 h2
Ih
M
AM
L(M
2) S
iste
r C
y/'T
BI
1st C
R
fl 83
1
9F
A
ML
(M7
J B
roth
er
Cy T
BI
2nd
CR
1 ha
plot
ype
mat
ch
CSA
CSA
CSA
CSA
CSA
C'SA
b1T
S
GrH
D
Die
d G
vHD
. No
evid
ence
of
leuk
aem
ia a
t p.
m.
BM: C
R D
ied
GvH
D. N
o ev
iden
ce o
f leu
kaem
ia a
t p
.m.
BM: C
R W
ell.
No
GvH
D
Rem
ains
wel
l and
in C
R
Rem
ains
wel
l and
in C
R
BM: C
R.P
B: n
o ab
norm
al c
ells
B
last
s ap
pear
in P
B BM
: 60
% b
last
s (M
2)
Die
d du
ring
rei
nduc
tion
\Vei
l. PB
: Nor
mal
R
ash.
PB:
Nor
mal
B
M:
10%
bla
sts.
PB:
Y',!;,
blas
ts 1
\111
D
ied f
ollo
win
g ch
emot
hera
py
7 8 9 10
11
12
I40
143 40
72
105
21
17
F
AML(M1 B
roth
er
1st C
R
14 M
c-
AIL
Si
ster
2n
d CR
23 F
A
MI.
iM31
Bro
ther
2n
d CR
44 h
l A
ML
lhl4
J Si
ster
1s
t CR
30 F
A
ML
(M4J
Bro
ther
1s
t CR
1
hapl
otyp
e m
atch
i M
c-
ALL
Si
ster
2n
d re
laps
e
Cyj
TBI
M
Cy/
TBI
Cy/
TBI
Cy/
TBI
CyJ
TBI
CSA
CSA
CSA
CSA
CSA
/MTX
MTX
24
30
31
33
2 4 6 9 10
I6
17
19
6 13
17
26
13
7 I3
8 14
0 7 - 13
31
8 74
81
90
25
31
36
37
51
Wel
l. PB
: Nor
mal
BM
: 75
% b
last
s BM
: 40% b
last
s. P
B: n
o bl
asts
(M
2)
Die
d du
ring
rei
nduc
tion
Wel
l. PB
: Nor
mal
BM
: CR
BM:
70%
blas
ts.P
B: 7%, b
last
s ic
-ALL
) BM
: 80
% b
last
s. D
ied
duri
ng re
indu
ctio
n
HM: r
elap
se
BM:
SO%
, bla
sts.
PB
: lW
%, bl
asts
(M3
) D
ied
duri
ng re
indu
ctio
n
BM:
3 3'
g bl
asts
D
ied i
n re
mis
sion
fol
low
ing
regr
aft
HM:
70%
blas
ts (
M4 1
BM
: 80
% b
last
s. P
B: 1
6'%
, bla
sts
Furt
her
ther
apy
refu
sed.
die
d
Bm:
100%
bla
sts.
PB
80%
bla
sts
(c-A
LL
) BM
: 7%
bla
sts
(c-A
IL)
BM:
8% bl
asts
(c-
ALL
) D
ied
follo
win
g fu
rthe
r re
laps
e
Abb
revi
atio
ns:
AM
L=a
cute
mye
loid
leu
kaem
ia (
FAB
type
in p
aren
thes
es):
ALL
= ac
ute
lym
phob
last
ic le
ukae
mia
. B=
B c
ell t
ype,
c =
'com
mon
' ty
pe: C
R =c
ompl
ete
rem
issi
on:
Cy =
cycl
opho
spha
mid
e:
TBI =
tota
l bo
dy
irra
diat
ion:
M
=m
elph
alan
: CS
A =
cycl
ospo
rin-
A:
MTX
=m
etho
trex
ate:
G
vHIl
=gra
ft
vers
us
host
di
seas
e: B
M =
bone
m
arro
w:
1% =
peri
pher
al b
lood
. C
hrom
osom
e re
sults
: -=
no
sam
ple.
Per
iphe
ral
bloo
d +
mito
gen
PHA
unl
ess
othe
rwis
e st
ated
(Il
n = n
o m
itoge
n. P
WM
=po
kew
eed
mito
gen.
LPS
=lip
opol
ysac
char
ide,
PH
A =
phyt
ohae
mag
glut
inii
i): Y
=Y
bodi
es i
n in
terp
hase
nuc
lei.
4 3 6
‘i’able I I . C’ytogenctic studies in Case 7. I3MY No. 140. Fcm:rlc paticnt / tiiiilc donor. I3onc marrow 3 1 weeks post 13M’I’.
No. of mctsphases
44 49 55 5 0 - 5 h . Culture + mar* 4h.W 4 h . X X + mar* + mar* + mar*
* Six marker chromosomes (thrcc of which showcd similar morphology) wcrc present in seven of the 1 3 ancuploid cells. and il 3q + chromosome in tivc cells. These abnormal chromosomes could not dctinitcly be identitied in all the ancuploid cells.
olall the metaphases of the marrow examined 3 1 weeks after HM‘I’, when it contained 40(%, of blasts, are shown in Table 11. Because a n aneuploid clone (Pig 1 ) with marker chromosomes was identified, it is most prohable that this represents a leukaemic cell population. I3onc miirrow was not available to us at diagnosis. but no cells belonging to the clone werc itlentitied prior to bone marrow transplantation.
Case 8. a male patient, is of particular interest, because only normal donor cells could be identitied chromosomally at relapse, although the interphase cells were predominantly ol recipient type. As can be seen from Fig 2 . the recipient cells occurred in aggregates and were identitied by a Y body, so we have concluded that these belong to the leukaemic clone. A Y chromosome was also identified in poor quality metaphases whose chromosomes could not even be counted. No cells with a n abnormal karyotype were detected prior to BM‘I’.
I n the case of the female patient No. 9, marrow was not available at diagnosis, but prior to transplantation no chromosomal abnormality was detected. For at least 2 0 weeks alter gralting the only cells found in division in the marrow were of donor origin. At the time of relapse the marrow, as judged by the virtual absence of Y bodies. was populated by recipient cells. whereas the cells in division i n the blood were of donor origin.
‘I’he data on the male patient No. 10 are scanty. His marrow was not available at diagnosis and yielded no mitoses when examined prior to transplantation. Kesults showed that the cells of the marrow reverted to recipient sex at the time of relapse.
‘I’he detililed cytogenetic tindings in the marrow at the time of relapse in Case 1 1 itre sho\vn in ‘I’able 111. It can be seen that there was a mixture of donor and recipient ~ ~ 4 s . N o normal recipient cells were found at any time and the pseudodiploid cells were only detected after the marrow had been cultured for at least 24 h. The best yield of the pseudodiploid cells
437
Fig 1 . Case 7 . Representative metaphasc with 55 chromosomes. Arrows indicate the 3q+ and marker chromosomes.
Fig 2. Case 8. Y-body in interphases and a Y chromosome in a mctaphasc in a group ofrecipienl cells.
438 Sylviu D. Lawlor et ml
Table Ill. Cytogenetic studies in Case 11. BMT No. 1 0 5 . Female patient/male donor.
No. of metaphascs
Pseudodiploid XX
Culture 4 6 , X Y 4 6 , X X Clone A Clone R
Bone marrow 74 weeks post HMT Direct 12 0 0 24 h 4 0 0 72 h 0 0 2 Total no. of cells 16 0 2
24 h 2 0 0 48 h 1 0 0 72 h 0 0 0
Total no. of cclls 3 0 0
Hone marrow 8 1 weeks post HMT
0 1 7
8
0 4 1
5
Fig 3(a). Casc 11. Clone A: Pseudodiploid with rearrangements.
439
Fig 3(b). Case 1 1 . Clone H: I’seudodiploid with rearraiigc.ments and two marker chromosomes.
was obtained in a 72 h culture. If only direct preparations had been examined. the pseudodiploid cell population would have escaped detection. Two pseudodiploid clones were detected in the first sample: clone A showed a translocation between the long arm of a No. 2 chromosome and the short arm of a No. 12 . and the addition of‘ unidentified chromosomal material to the short arm of a No. 2 1 : clone B showed the loss of a No. 1 3 and a No. 1 5 chromosome, the gain of two small subtelocentric marker chromosomes and the presence of extra chromosomal material of uncertain origin on the long arm of a No. 3 chromosome. Only the clone B was found in the later bone marrow sample (Fig 3 ) .
Case 12 is included because the marrow showed only donor cells in division at a time when 7% blasts were present following reinduction of partial remission after relapse. Unfortunately when the percentage of blast cells was 1 OO‘%,, no mitoses were obtained. This patient was one of the earliest to be investigated and at that time we were not aware of the importance ofdefining the status of cells in interphase. I t is interesting to note than when the blasts were present in the blood. the PHA-stimulated cultures yielded only cells of donor origin.
D I S C’ LJS S I 0 N
These studies, using the sex chromosomes as markers of cellular origin following allogeneic bone marrow engraftment. have shown that when the procedure is successful, donor cells populate both the myeloid and lymphoid series. This outcome of bone marrow transplan-
440 Sylvia D. Lawler et a1
Table IV. The origin of myelod and lymphoid cells following bone marrow transplantation
1,eukaemic type ALL or AMI,. AMI, or A I L AMI,. AMI,.
- - - - -
No leukaemic Leukaemic Leukaemic Leukaemic relapse relapse relapse relapse
Clinical features Graft persists. Normal donor cells in division in bone marrow. Lymphoid cells donor. No Kecipient cells.
Pattern I
Graft persists. Graft persists Graft rejected. Marrow but no donor No donor cells. chimaerism. cells in marrow. Kecipient cells Lymphoid leukaemic. cells donor. Lymphoid cells donor.
II 111 IV
tation is designated as Pattern I in Table IV. Early marrow engraftment is shown by rising levels of granulocytes by 2 weeks after the marrow has been transplanted. This success is mirrored by the presence of donor metaphases in the marrow. As far as lymphoid cells are concerned, the successful graft is associated with the presence of donor metaphases in peripheral blood cultures, whether the mitogen used stimulates predominantly the T o r H cell class of lymphocyte.
In this series, host cells that had not been eliminated or chromosomally damaged by the chemotherapy and/or TBI that preceded the grafting. were found in three cases who did not have leukaemic relapses.
Case 4 is exceptional because the patient remains well and in complete remission from B-cell acute lymphoblastic leukaemia, even though karyotypically normal recipient cells were found on several occasions in blood cultures stimulated with 1,PS and PWM.
Our studies in the patients who have relapsed (Cases 5-12) show that relapse following BMT is associated with various chimaeric states in marrow and blood (Table IV). In Pattern 11, at the time of relapse, there is a mixture of normal donor cells and recipient cells in division in the marrow. This suggests that, for a time, the leukaemic population is contained by competition with the normal cells. Whilst this is happening in the marrow, the peripheral blood lymphoid population remains of donor origin. Any recipient elements identified a t interphase or metaphase in the peripheral blood are probably formed myeloid elements in interphase, or leukaemic blasts in mitotic cycle that have spilled over into the circulation.
Cytogenetic Studies in Allogcneic BMT 44 1
Competition between normal bone marrow cells and leukaemic cells must be a stage in the evolution of all types of leukaemia. Our study of the events in the marrow following BMT has enabled us to demonstrate this situation. Another factor that must be considered is that the stromal cells of the marrow in successful engraftments have been shown to be of donor type (Keating et al, 1982).
When all the normal cells in the marrow have been replaced by those of the recipient, the relapse pattern is of Type 111. peripheral blood lymphoid cells still being of donor origin. Using HLA identical sibs as donors, we have not yet found a patient in leukaemic relapse following BMT who does not still have predominantly donor T cells. This is also the case in patient No. 11 who received a BMT from a brother matched by one haplotype.
The fact that the circulating lymphoid population remains of donor type during relapse could be due to the life span of some T lymphocytes. Leukaemic relapse may cause death of the patient before host lymphoid cells can repopulate the blood. The observation that the marrow can contain recipient cells whilst the blood lymphoid cells are ofdonor origin is an illustration of the separate origin of the myeloid and T-cell lineages as, for example, is well established in cases showing the Philadelphia (Ph' ) chromosome. In chronic myeloid leukaemia the Ph' chromosome is regarded as a marker of the stem cell precursors which give rise to platelets and granulocytes whilst the progenitor cells giving rise to the T cells are not usually marked by this translocation (Lawler. 1977).
Pattern number IV, in which all myeloid and lymphoid elements are of recipient type, shows that the graft has been completely rejected. This pattern applied to Case 6, who initially had a successful take of the marrow graft. Absence of donor lymphocytes provided early evidence of relapse in this patient, who received marrow from a brother matched by one haplotype.
In this series we have found in the marrow, following BMT for ALL, normal donor cells in division during relapse, whilst the presumed leukaemic cells were of recipient origin but could only be identified by the presence of a Y body. Undoubtedly we would have concluded that in the male patient No. 8, the leukaemic relapse had occurred in the cells of the donor, if we had not identified Y bodies among the cells in interphase and unanalysable metaphases. A study of this case illustrates a point that is relevant to cytogenetic investigation of bone marrow in leukaemia at any time. Cells with a normal karyotype in a sample of bone marrow from a patient with leukaemia do not necessarily belong to the leukaemic cell population.
In our series we have not yet found evidence of relapses occurring in cells of the donor, although such cases have been recorded (Fialkow et ul, 1971: Thomas et aJ, 1972: Goh & Klemperer, 1977; Elfenbein et al, 1978; Newburger et al. 198 1 ). The possibility exists that in some of these cases the patients could have had a mixed population of cells in the marrow at the time of relapse, so that the observed karyotype represented dividing donor cells and the non-dividing population was of the leukaemic recipient type. On the other hand, Thomas et al ( 1972) found Y bodies in 50% of lymphoblasts and XY metaphases in the marrow of a female in relapse after BMT from a male donor. The case described by Elfenbein ef al (1978) is complicated, depending on a single autosomal polymorphism to identify the donor origin of an abnormal clone in the marrow carrying a translocation t( 8;2 1 ), that is characteristic of FAB-type M2, but in this case the cells of the clone had one X chromosome and no Y and
442 Sylvia D. Liiwlrr r t a1
therefore could not be sexed. After HMT the patient also had a chloroma, which on the basis of the presence of Y bodies in interphase, was identitied as being of host origin.
Newburger ~t a/ (1 98 1 ) described a patient with acute lymphoblastic leukaemia who relapsed following a BMT which was preceded by high dose cyclophosphamide and mi. The donor origin of the leukaemic cells in relapse was established by the use of a n autosomal marker. The antigenic phenotype of the leukaemia was shown to correspond with that ofthe original disease.
We were not able to relate the karyotype of the metaphases found in relapse with that seen at diagnosis because of lack of information. Aneuploid cells with chromosome numbers as high as 5 5 (Case 7) are certainly very unusual in acute myeloid leukaemia de m v o . In Case 1 1 the two pseudodiploid clones seen in relapse had unbalanced karyotypes and the type of chromosomal rearrangements that could result from the effects of irradiation. It is possible that both the clones arose from the original leukaemic cells and that they competed with each other and the normal donor cells and eventually one clone became predominant.
The purpose of the BMT procedure is to ablate the leukaemic state. It is clear that this is not always achieved and that recipient cells of leukaemic type can repopulate the marrow. Occasionally leukaemic transformation of donor cells has been demonstrated and various explanations for this have been suggested. A leukaemic agent in the recipient may be capable of activating a latent oncogenic virus which persists after RMT. Alternatively, patients are prone to viral infections after transplantation. and in the presence of this type of virus, fusion may take place between donor and recipient cells. However, according to our experience and that of Boyd et a ] ( 1982), the majority of relapses after BMT occur in the cells of the recipient.
A C K N 0 W I . F. I)(; M K N 'I'S
We thank Dr D. G. Spence and Dr M. J. W. Faed for information about Case 7, Dr 13. K. Keeves for a constructive criticism of the manuscript, the Photographic Department of the Koyd Marsden Hospital for the illustrations and Miss M. P. Leach for typing the manuscript.
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