Biologic and Cytogenetic Characteristics of Leukemia in Infants

BATIA STARK, MD,* RUTH VOGEL, MD," IAN J. COHEN, MD,* TEHILA UMIEL, PHD,* ZIPPORA MAMMON, MSc,t GIDEON RECHAVI, MD, PHD,* CHAIM KAPLINSKY, MD,* DANIEL POTAZNIK, MD," AMALIA DVIR, MD,' YITZHAK YANIV, MD," YAKOV GOSHEN, MD,' NURlT KATZIR, PHD,* BRACHA RAMOT, MD,* AND RlNA ZAIZOV. MD'

Clinical features, leukemic cell characterization, chromosomal findings, and treatment outcome were analyzed in a retrospective study of 30 cases with acute leukemia of infancy, 24 infants with acute lymphoblastic leukemia (ALL), and six cases with acute nonlymphoblastic leukemia (ANLL). Extensive bulky disease with organomegaly, central nervous system (CNS), and skin involvement were prominent features at diagnosis with a higher frequency in ANLL as compared to ALL. Four of six ANLL patients were classified as monocytic or myelomonocytic. In the ALL group nine of 24 (36%) were non-L1 morphology and six of 17 (33%) were common ALL antigen (CALLA) negative, the majority of them (five of six) were included in the non-Ll group. Immunophenotyping revealed four cases with early B-cell (three patients: Ia+B4+, and one patient: Ia+) and two cases with T-cell. Mixed lineage leukemia was found in five infants. Heavy chain immunoglobulin gene rearrangement was present in six cases tested, two CALLA+, two with Ia+B4+, and two were undifferentiated mixed lineage leukemia. Chromosomal aberrations were detected in ten of 18 patients, mostly in ANLL and CALLA negative ALL. Translocations were detected in six patients, involving 4q21-23 and l l q 2 3 in three and two cases, respectively. The probability of five-year DFS was 27% for the whole group. The worst prognosis was observed in infants younger than 6 months of age, in whom the leukemia cell characteristics was compatible with stem cell: ANLL, very early pre-B, or undifferentiated mixed type. The chromosomal aberrations found in all cases included translocation with the seemingly nonrandom breakpoints at 4q21 and 1lq23, and breakpoints that corresponded to known fragile sites. This finding may be suggestive of an underlying genetic predisposition associated with the poor prognosis of leukemia of infancy. Cancer 63:117-125, 1989.

and myeloid leukemias are for treatment selection. In contrast to the high cure rate achieved in children with ALL,8 a much lower rate is clinically and biologically heterogeneous diseases reported in infants less than 1 year of age (60% verfor which specific therapeutic regimens should be desus 20%).9-12 signed.1-4 In acute nonlymphoblastic leukemia (ANLL) as a conIn acute lymphoblastic leukemia (ALL) numerous factors were found to have a prognostic ~ignificance,~-~ sequence of the overall poor outcome, the prognostic significance of biologic parameters remains equi~ocal.~.' among which age and initial leukocyte count are the major Nevertheless, the young age seem to fare worse than the inldicators of outcome and are used for stratifying patients older children (20% versus 50% three-year continuous complete remission [CCR]).4 Presented in part at the SIOP meeting, Belgrad, Yugoslavia, 1986. The young age adverse effect may be partially explained From the *Sambur Center for Pediatric Hematology Oncology and the ?Laboratory of Clinical Cytogenetics, Beilinson Medical Center, Peby the segregation of known high-risk factors such as tach Tikvah, and the $Hematology Department, Sheba Medical Center, high and extensive leukemic blast burden at presentaTel Hashomer, Sackler School of Medicine, Tel Aviv University, Tel t i ~ n , ' - ' ~morphology "~ of myelomonocytic or monocytic Aviv, Israel. The authors thank Pearl Lilos of the Statistics Laboratory, Tel Aviv type,14-16and high incidence of negative common ALL University, for the statistical evaluation of the results, and Emanuel Nir, antigen (CALLA) phenotype. l 7 Some investigators attribPhD, Kaplan Hospital, Rehovot, Israel, for the election microscopy inuted the poor outcome to excessive toxicity and subopveirtigation. Address for reprints: Batia Stark, MD, Sambur Center for Pediatric timal therapy," whereas others disagree with these obHematology Oncology, Beilinson Medical Center, Petach Tikvah, 49 100, servations. Intensive regimens introduced recently Israel. may lead to improved result^.^^-^^ Host factors influencing Accepted for publication July 11, 1988.
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the milieu of the blasts or drug metabolism cannot be ruled out and require appropriate analysis with fully matched older control patients. A retrospective study is reported, analyzing clinical and biologic characteristics of acuted leukemia in infants in whom the malignant clone is unique and characterized by a clone of stem cell origin and nonrandom chromosomal translocations. The features may be suggestive of a special genetic event underlying the malignant transformation,which may contribute to the poor prognosis. Materials and Methods

American-British (FAB) morphologic criteria25were assigned in 18 patients, whereas in the previous patients classification was limited to either lymphoid (L) or myeloid (M) morphology. Cytohistochemical staining including periodic-acid-schiff (PAS), acid phophatase (ACPH), myeloperoxidase (MPO), Sudan black (SB), chloracetate esterase (CAE), and nonspecific esterases (NSE) were performed in 23 patients using standard technique. Definition of the leukemia type for treatment assignment was based on the morphology on light microscopy and cytohistochemistry.

Patients and Treatment

This retrospective study material comprised 30 infants younger than 2 years of age diagnosed with acute leukemia, admitted to the Sambur Center of Pediatric Hematology Oncology, Beilinson Hospital from 1966 to 1986. The group represents all leukemic patients under the age of two years treated at this center. Ten patients were younger than 1 year of age, six of them were younger than 6 months of age. During the 20 years of the study period, different therapy regimens were used, which can be roughly divided into three main treatment categories: 1. The ANLL protocol (six patients) using induction with cytozin arabinoside (CZ), rubidomycin (RB), 6thioguanin (6-TG), cyclophosphamide (CTX), and vincristin (VCR) and monthly pulses with lower doses of CZ, RD, and 6-T6. 2. The acute leukemia protocol used from 1966to 1969 (four patients), which included induction of 6-mercaptopurine (6MP), and prednisone (P) and monthly pulses with VCR,P, and oral therapy with 6MP and MTX with no CNS prophylaxis. 3. Acute lymphoblastic leukemia (ALL) protocol during 197 1- 1986 (2 1 patients) using induction with VCR,P, with the addition of either CTX (two patients), or CTX and adriamycin (AD) (six patients) or CTX, AD, and CZ ( 18 patients). Consolidation included asparginase (ASP) (1 8 patients), occasionally with high-dose IV MTX (three patients) and in the last two years an additional four pulses with VCR, MTX, ASP (seven patients).24Maintenance pulses were given six monthly and then an additional six bimonthly including VCR, CTX, AD, CZ, and P. Central nervous system (CNS) prophylaxis comprised of IT MTX (five patients) or IT triple with MTX, CZ, P ( 16 patients). Cranial irradiation was given to seven infants older than 9 months of age, 1500 to 2400 cGy.

Immunologic Characterization
Immunologic characterization was performed in 20 patients.27328 A panel of monoclonal antibodies defining T-cell-restricted antigens: CD7-3A 1,29 CD2-T 1 1, CD30KT330; B-cell restricted antigens: CD 19-B4,3 CD20B 1,32 CD2 1-B233; myeloid restricted antigens: CD 13MY7,8,34CD33-My9, C D W ~ ~ - M Y and ~, B-cell ~ ~ ; associated antigens: HLA-DR-Ia36 and CD 10-J5 CALLA37 were utilized to determine the phenotype of the leukemia cells. Cell surface antigens were analyzed by indirect immunofluoresence or flow cytofluorometry as previously de~cribed.~.~~

Immunoglobulin Gene Analysis

DNA was extracted from leukemic blasts, digested by the restrictive enzyme ECO-RI and analyzed by Southern blot hybridi~ation~~ with immunoglobulin heavy chain J region radioactive probe (kindly donated by Dr. P. Leder) in six patients.

Karyotyping
BM aspirate cells and peripheral blood blasts were processed both directly and after methotrexate synchronized culture in 18 patients. Air dried preparations were stored for a few days and routinely stained for G bands by the method of Seabright.39 In four patients only conventional Giemsa staining was performed.

Statistical Analysis
Disease-free survival (DFS) was estimated with the use of Kaplan-Meier methods. The notable event was failure to achieve remission and relapse (death during remission occurred only in one patient late in the course of the disease). To test the quality of survival curves we used generalized savage (Mantle-Cox) test.40 Pearson chi-square was used to measure the association between variables. In cases of 2 X 2 tables, Fisher exact test was used.40

Morphologic and Cytochemical Studies

Wright Giemsa stain morphology of bone marrow (BM) aspiration was examined in all patients. The French-

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119

TABLE 1. Clinical and Laboratory Characteristics of Leukemic Infants (1966-1977) I* 2 6 O/F 6 ND 39 L2 ND ND ND BMSK 0.5 3
18

4 3 A/F 10 ND 18 M4 ND ND ND BM 0

6 9 A/F 3
-

7 24 A/F 0
-

8 19 AIM 0
-

10

Age (mo) Origin/sex Liver (cm) CNS Leukocyte count (X lo9 1) Morphology (FAB) PA:S NS E MF'O 1st relapse CCR (mo)

21 O/M 5 ND 7 L ND ND ND BM 8

O/F 6 ND 70 L2 ND ND ND BM 8

4.5 AIM 5
-

15 BIM 6

+
-

19 L ND ND ND CN 12

148 L1/2 ND ND ND
-

25 LI ND ND ND
~

20 M3 ND ND

139 M4

+ + +

350 L

ND
-

140t

110+

BM 5

BM 0

BM 11

N D not done; M: male; F female; 0: Oriental Jew; A: Ashkenazi Jew; B: Beduin Arab; L: lymphoid; M: myeloid; BM: bone marrow; S K skim; CNS: central nervous system; CCR: duration of continuous com-

plete remission; +: alive in remission. * Patient nos.

Results

Clinical and Laboratory Features at Diagnosis

Among the 30 evaluated leukemic patients younger than 2 years of age the female/male ratio was 15/15. Female predominance (F/M of 7/3) occurred under the age of 1 year, whereas male predominance (M/F of 13/7) occurred at the age of 1 to 2 years (Tables 1 and 2). Ethnic origin was distributed between ashkenazi (1 5 patients) and oriental Jews (12 patients) or mixed (two patients), with only one patient of Beduin extraction, and it was similar to that observed in the older childhood leukemic population. Among the very young infants (younger than 6 months of age) five of six patients were of askenazi jewish extraction. Prenatal history included maternal medication with thyroxin, antihypertensive drugs, intramuscular iron injection during pregnancy, and paternal medication with anti-psychotic drugs, one of each. Three patients had healthy twin siblings, one had a brother with Down syndrome, and another one had a sister with heterochromia. Physical abnormalities in the patients included one with horseshoe kidney and one with heterochromia. Clinical findings at presentation of hepatomegaly, spllenomegaly (>3 cm below the costal margin), and CNS involvement occurred in 50%, 56%, and 20% of the patients, respectively. The incidence of CNS involvement w,as significantly higher than in the older population (aged 2 to 17 years, 4.1%; P < 0.01). A higher frequency of hepatosplenomegaly 1OO%, and CNS involvement 50%, although statistically not significant, was observed in the younger subgroup (younger than 6 months). In one patient (no. 20) leukemic CNS involvement preceded by a few w'eeks BM involvement. Subcutaneous infiltrations were found in three of six very young (younger than 6 months) patients, (nos. 2,2 1, and 22) and in one of 24 of the older ones (no. 3). In two infants it preceded peripheral blood

dissemination by 12 and 3 weeks (nos. 2 and 22). High leukocyte count > 100,000 cells/mm3 was presented in 20% of the patients, low hemoglobin level <6 g/dl in 46%, and low platelet count of t20,000/mm3 in 30%. There were no significant differences in these parameters in the two age subsets (younger than or older than 6 months), and in the older population (younger than or older than 2 years).

Leukemic Blast Cell Morphology

Leukemic blast cell morphology defined by light microscopy and cytohistochemistry was lymphoid in 80% and myeloid in 20%. In the lymphoid type out of 24 patients 12 had L1 morphology, seven L1/2 or L2, and two had L1/3 morphology; three of the 24 were defined retrospectively as L1 (Tables 1 and 2). In two very young infants (nos. 23 and 24), ultrastructural morphology was of undifferentiated leukemia and in the last one with cytoplasmic microfibrilis situated in the nuclear identation and diffuse alpha-D glucoronidase stain in light microscopy, suggestive of a myelomonocytic origin. Among the six myeloid leukemic cases the FAB morphology distribution revealed M5 in two cases, M4 in two cases, M3 in one infant, and in the sixth infant (no. 21) the morphology was indistinguishable between myeloid or L2. The blast cells were stained with PAS positive and weakly NSE. In electron microscope morphology was of undifferentiated blast with weakly positive ultrastructural myeloperoxidase reaction.

Phenotypic Characterization
Phenotypic characterization was performed in 20 patients (Table 2) in one by E,EA rosseting and the rest by monoclonal antibodies. Ia+ B4+ CALLA+ B1- B2phenotype was encountered in ten of the 16 (62.5%) of the examined ALL patients. Among the six CALLA neg-

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: :
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ative cases, two patients had T-cell phenotype (T 1 1+), three patients had Ia+ B4+ CALLA-, compatible with the earliest recognizable phenotypic stage of B-cell differentiation, and one patient had undifferentiated Ia+ blasts. Three patients had myelomonocytic phenotype. Mixed phenotypes, manifested by overlapping of antigenic expression of myeloid and lymphoid antigens, were found in five patients. In two young infants with undifferentiated morphology (nos. 2 1 and 24), the first one revealed a phenotype of Ia+ My9+ My7+ and B4+ by double staining technique,27328 and in the second patient with only Ia+ at presentation, B4 antigen was acquired during relapse and My7 antigen in in vitro culture. Two other patients with morphologic defined ALL CALLA+ (nos. 28 and 29) coexpressed My4 antigen and additional patient with monocytic leukemia (no. 20) coexpressed My9+ My7+ My4+ and B4+ antigen.

Heavy Chain Immunoglobulin Gene Analysis

N

r N

This analysis showed in all six examined patients rearranged genes in addition to one germ line allele. It included two cases with CALLA positive ALL, two with early Bcell phenotype Ia+ B4+, and the two infants with the undifferentiated mixed leukemia.

N 0

Karyotype
0 '

Evaluable cytogenetic analyses were performed in 14 patients (Table 3). Abnormal karyotype was identified in ten cases and included all examined (six of six) infants younger than 7 months, the majority of the ANLL patients (five of six), and the CALLA negative ALL patients (five of six). Aberration could not be exactly defined in three patients in whom only conventional Giemsa staining was performed. In one patient with T-cell leukemia 6q- was found during relapse, and in six other patients translocations could be identified.

Response to Therapy and Complications

Complete remission (CR) was achieved in 27 of 30 patients (90%). The ANLL and the younger subset fared worse than the ALL group and older subsets, attaining complete remission of 50% and 66% versus 100% and 95%, respectively. The patients who failed to enter remission died five to 16 weeks from diagnosis. Outcome of the patients is summarized in Table 4. Of the 27 patients who entered complete remission, 16 relapsed, of whom 14 died. Additionally, one patient died during remission from a secondary tumor. CNS relapses occurred in seven patients, in four of them as an isolated event. They included two young ANLL patients with CNS involvement at presentation and the other five patients with ALL, two of whom with T ALL. No significant risk

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TABLE 3. Cytogenetic Findings

121

factors for CNS relapse could be identified including the various CNS prophylaxis regimens. The estimated probabiility of DFS for all leukemic infants was 27% at five years, with median follow-up of 36 months. The most significant poor prognostic factor found by univariate an,alysiswas age (younger than 6 months) and ANLL type of leukemia with close association between them. One year DFS for the ANLL and the very young group was 0741,whereas that of the ALL and the older infants was 70% and 65%, respectively (P < 0.00001) (Figs. 1A and B). The very young infants comprised a unique group with a high incidence of known poor prognostic characteristics such as ANLL (four of six), non-L 1 CALLA negative ALL (two of two), and chromosomal translocations (three of three). Of the older infants the two ANLL patients and the ALL non-L1, CALLA negative patients all relapsed within 39 months from diagnosis, whereas the fiveyear DFS of ALL L1 and CALLA+ was 70% and 50%, respectively. The three patients with initial CNS involvement also fared poorly and died within one year of diagnosis. Severe late sequelae occurred in two patients (nos. 6 and 7), who were diagnosed at the age of 9 and 24 months, and received cranial irradiation of 1500 and 2400 cGy, respectively, and IT MTX. Both developed meningomas five and six years following radiation to the skull. In the first patient meninginoma was resected and recently recurred. She is alive in remission for 12 years from diagnosis with moderate brain damage. In the second patient a malignant transformation to meningosarcoma occurred and she died three years later, nine years in continuous complete remi~sion.~'

Patient
no.

% ABN

Source PB BM BM BM BM MX BM PB BM BM BM BM BM BM PB BM

Status

cells
II 100 65 81 100

Karyotype

4 9
14

D
RI RI D D

19 20
21'

D
D RI RI RI RI RI RI RI D

I00 LOO
45

36
19

22

0
21 14

23'
24

80 0
85 75

30

46XX-A-6+MI+M2 t(l:21)? 45XY-A+small metacentric M tril9? 46XY 6q47XY +t( I :3)(q,,:P I4)? 46XYt(ll:14)(q13-21:q24 or 32) 46XXt(I I:14)(q13-21:q24 or32) 46XYt(I I:17)(q23:P13) 46XYt( I I:17)(q23:p13) 46XY 46XYt( I I:17)(q23:P13) 47XYtri8:t(l 1:17) 46XX 46XXt(4:7)(q2 I :P 13)I7P+ 45XXt(4:7)(q2I:PI 3)- I7P+ 47XXtri8 46XXt(4:I l)(q21:q23) 46XY 46XYde14q23-28:19q+t(4: 19)? 47XY+M tril7?

ABN: abnormal: PB: peripheral blood; BM: bone marrow: MX: maxilla: M: marker chromosome. Status: D, diagnosis: RI, relapse. * Patients 21 and 23 were previously reported2'.**.'*.

being significantly more frequent' are either an independent variable of poor prognosis44or associated with other poor risk features.45The distribution of myeloid leukemia differs in infants as compared to older children, with a higher prevalence of the poor prognostic subtypes, the myelomonocytic M4, and the monocytic M5.4In the current study four of six infants exhibited M4-M5, one M3, and one undifferentiated mixed subtype.

Discussion Leukemia in early infancy has a most grave outlo,,k.4,9-12,i9-21,42,43 I n our series the five-year DFS was 32% for ALL younger than the age of 2 years. For ANLL only 50% achieved remission and this lasted no more than five months. Similar results were reported in ALL and ANLL by other investigator^.^. lo-' 2*1 The role of the various parameters contributing to the poor outcome in acute leukemia in infancy is not as yet clearly defined. The infants with ALL in our series and as reported by others exhibited bulky and extensive disease at presentation, along with higher incidence of hepatosplenomegaly, CNS involvement, high leukocyte counts, lower platelet count, and lower Hb level^.^-'^ In ANLL extensive disease was even more pronounced with a high frequency of extreme leukocytosis and extramedullary involvement of skin and CNS.',14,16
'3

TABLE 4. Outcome of Leukemic Infants Related to Age Group and Subtype of Leukemia
0-24 mo
0-24 mo ANLL+ALL (30)
Slow responders* Induction failure Entered remission Death in remission Relapses BM BMICNS CNS Test BM/Skin Live in remission On treatment Off treatment

0-6 mo

7-241110

(6)

(24)

ANLL (6)

ALL (24)

6/27

3
27

It
7 3 4
1 1

214 2 4 0

4/23 I 23

313
3 3
0

It
1

3/24 0 24

It

I
2 0

6
4

I
2 0

6
I
4 1 1

0
1

0
8

0
0

8$

48

0 0

8 4

Morphology The morphology of the leukemic cell in infants differs from that of older children. The non-L1 lymphoblasts

Results are expressed in number of patients; number of patients in parentheses. as those infants who did not attain complete remission by 28 days. t The patient (no. 7) died nine years from diagnosis of secondary meningiosarcoma. $ All patients are in first complete remission except for patient no. 27 who is in second remission after CNS relapse. 5 Patients are in first remission 65, 104, and 140 months from diagnosis. One patient (no. 16) is in second remission two years after BM and CNS relapse, following bone marrow transplantation.

* Slow responders were defined

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-----Age-7-24 mo(N=24)

211

01

i i

28

'

42

'

& ' m

84

s s

112

126

140

TIME (Months)

FIGS.1A AND 1B. The estimated probability of DFS curves estimated by the Kaplan-Meier methods. Relapse or failure to achieve complete remission were included in the definition of the disease. (A, top) DFS related to age at diagnosis. (B, bottom) DFS in infants older than 6 months (7 to 24 months) related to subtype of leukemia and FAB morphology.

Immunologic Phenotype We and others reported a low incidence of both CALLA+ lymphoblasts and T-cell phenotypes in infants compared to older children.'.' '3'2-'7,46347 In four patients, early null "pre-pre-By cell phenotype Ia+ B4+ CALLAwas encountered. This is the most frequent leukemic subtype reported in i n f a n ~ y ' * ' ' * ' ~ and ,~~ represents ,~~ the earliest phenotypically identifiable stage of the B-cell lineage. At this early stage evidence for B-cell commitment at the DNA level is provided by the uniform presence of immunoglobin heavy chain gene rearrangement, retaining the germ line light chain gene,46.48-5' as found in the two infants with Ia+ B4+ leukemia. Recent data5' demonstrated Ig heavy chain without light chain gene rearrangement providing further evidence for an earlier stage of Bcell lineage in ALL of infancy. Undifferentiated leukemia is characterized by electron microscopy and immunophenotypically coexpressed features of mixed lymphoid myeloid origin in two babies. In both cases blasts could be induced to differentiateto either myeloid-macrophage, or to CALLA+ B-cell, depending on the growth factors administered in vitro. Immunoglobin heavy chain gene was also rearranged. Undifferentiated leukemia is a rare subgroup of acute leukemia, with

lineage defined by blast cell characterization Some of the undifferentiated Ia positive leukemias may correspond to the earliest stage of B-cell in which immunoglobulin heavy chain gene may have already been rearranged.48349 This subtype may be equivalent to the TdT positive acute lymphoblastic null cell leukemia, defined by Hoflbrand et al? Other subgroups may correspond to the early myeloid series.53Alternatively, those undifferentiated blasts may coexpress early markers of both lineages, indicating a multi-potential stem cell origin. As reported, examples were provided by the existence of myeloblasts positive for TdT lymphoid enzyme or/and with rearranged DJ region of the immunoglobulin heavy chain gene and T-cell receptor alpha chain gene.56 Those early markers and genetic steps are necessary but possibly insufficient for the lymphoid lineage commitment, and may reflect an immortalization of a transient promiscuous phase in a normal multi-potential prog e n i t ~ r . ~The '.~~ two babies with the early undifferentiated mixed characteristic leukemia may correspond to such a stem cell origin. Mixed leukemia in an otherwise more differentiated blast may reflect an abberrant gene expresExamples for this alternative sion (i.e. lineage ir~fidelity).~~ could be illustrated in three of the older infants, in whom the lymphoblastic CALLA+ leukemia expressed myeloid antigens and vice versa.56s9.60 The distribution of myeloid leukemia differs in infants as compared to older children, with a higher prevalence of the poor prognostic subtypes the myelomonocytic M4 and the monocytic M5.4 In the current study four of six infants exhibited M4-M5, one M3, and one undifferentiated mixed subtype. Cytogenetics Cytogenetic abnormalities were found in ten of 18 patients examined, in all infants under the age of 7 months. The six translocations identified included t( 1 1:17), t(4: 1 l), t(4:7), and t(4: 19) in four babies and t( 1:3) and t( 1 1: 14) in older infants. Nonrandom chromosomal aberrations are associated with specific types of leukemias, with certain morphologic features and/or with certain age groups indicating different and consistent etiologic mechanisms. The specific chromosomal aberration has been correlated with the course of the disease and progno~is.~'-~' In a recent report structural chromosomal aberrations were found most commonly in infants younger than 9 months of age, those involving the following chromosomes in order of frequency7*:1 lq, 9p, 16q, lop. 1 lq23 band seems to be the most frequently involved in infants leukemia and includes translocation with various chrom o s o m e ~ . ~The ~*~ leukemias ~ - ~ ~ associated with those translocations varies according to the recipient chromosome revealing a wide spectrum of morphologic and immunologic phenotypic feature^.^^,^^-^^ Translocations in-

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volving chromosome 11 were found in two patients, two weeks and five months old, t( 1 1: 17), t(4: 1 1). The t(4: 1 l)(q2 1:q23) was described in all ages but more frequently was reported either in infants (mainly in undifferentiated, mixed, or very early B-cell acute lymphoblastic leukemia (as in our study), or rarely was found in CALLA+ ALL and in ANLL.78379 The broad spectrum of phenotypic lineage provides evidence for a multi-potent stem cell in which the leukomogenic event involved 11~123. Chromosomal translocations involving 4q2 1 were found in three of four babies (younger than 7 months of age). In addition to the aforementioned pre-B-ALL with t(4: 1 l), the translocations found were t(4:7)(q2 1 :pl3) in myelomonocytic leukemia and t(4: 19)(q23-28:ql3) in an undifferentiated mixed leukemia; 4q involvement was also reported with a wide spectrum of leukemias such as t(4: 17) in unclassified leukemia with lymphoid and myeloid antigens and trisomy 4 in ANLL with multi-lineage involvement and in myelodysplastic syndrome.8 It is therefore suggested that 4q2 1 and 1 lq23 may be involved norirandomly in the transformation of a multi-potential progenitor. The recipient chromosomes for translocations of 4q or 1 lcl were 7p, 19q, and 17q. Translocation ( 1 1:17) involving 17q was found in our series in a newborn with mixed undifferentiated leukemia and was reported in adults with myeloid leukemia of all the FAB morphologic spectrum or with myelodysplastic Chromosome 19 was involved in our study in t(4:19) in an infant with Undifferentiated mixed leukemia. Aberrations involving chromosome 19q in t( 1 1:19) and t( 1:19) has been reported frequently in childhood leukemia with a wide range of phenotype^,^^*^^-^^ mainly pre-B-ALL with t( 1:19)67;7p was associated with myeloproliferative disorder in one infant, as reported in adultss3 Iroto-oncogenes located at various breakpoints have been demonstrated to play a role in the evolution of the malignant transformation. Proto-oncogene Hu-ets- 1 located on band 1 lq23 has been shown to translocate, rearrange, or amplify in human hematologic malignancies involving 1 1q23.66,76,84,85 In viral leukemia E26 Ets- 1 was shown to block differentiation in transformed myelob l a : ~ t s .The ~ ~ ,coding ~ ~ gene for protein p53 mapped to chromosome 1 7 ~ 1 3 was found to be overexpressed in lyniphoproliferative disorders where it is claimed to participate in the neoplastic proliferati~n.~~ Conversely, in myeloproliferative disease and ANLL including our patient, p53 was not expressed.87 Genes encoding for growth factors and their receptors have been mapped to chromosomal regions in the vicinity of ithe breakpoints mentioned above, for example, epidermal growth factor (EGF), T-cell growth factor (TCGFinterleukin-2) on 4q2 1,88 Erb-B oncogene, the closely related epidermal growth factor receptor (EGFR) on 7p13,

and the transforming growth factor beta type (T6FP) on 19q13.88 Autocrine growth regulation by inappropriate qualitative and quantitative alteration of growth factors and/or their receptors has been considered to play a primary and/or permissive role in c a r c i n o g e n e ~ i s The .~~~~~ role of growth factors has not been explored as yet in infant leukemia. Neoplastic transformation, a conditioned multi-step process that usually develops s10w1y,86-91-93 when occumng in a very early age may assign to an underlying genetic predisposition or other embryonic event, the first hit of tumorigenesis. Fragile sites, hereditary or constitutive, could serve a predisposing factor of such a first hit. These regions are susceptible to somatic recombinations and reside in proto-oncogenes and other active genes of cell diff e r e n t i a t i ~ n .In ~~ five . ~ of ~ six infant patients chromosomal translocations had remarkable concordance with the localization of the heritable (h-fra) and constitutive-common fragile (c-fra) sites: 1 lq23, 1 lq13, 1 7 ~ 1 27pl1, , and 14q24, respectively. Deletion or mutation of anti-oncogenes, defined as tumor suppressor or differentiation gene^,^^-^' could serve as another mechanism for the precipitation of a neoplastic transformation in the perinatal period. These events are usually associated with developmental malformations as observed in hereditary embryonal turn or^.^^-^^ Regulatory genes could reside on 1 lq23, the most frequently involved band in infant leukemia. This band is involved in malignant proliferation, developmental aberrations, or a combination of Among the malignant neoplasms, it was reported in hematopoietic and nonhematopoietic malignancies, such as Ewings sarcoma or neuroepithelioma with t( 1 1 :22).99 Developmental aberration was reported in constitutional chromosomal rearrangement t( 1 1:22).IooBoth processes combined are illustrated in the congenital leukemia with t(4: 1 1) and congenital In our patient with such translocation it was associated with heterochromia. Other supportive evidence is provided by the experimental setup of human cell hyrbrid (HeLa X fibroblast) where chromosome 11 was found to have a suppressor influence on tumor growth.Io2Investigation on the molecular levels may clarify the role of each factor and their intercorrelation in the evolution of infant leukemia. Chromosomal aberrations were found primarily in the very young infants. This finding is known as a poor prognostic marker. The most striking observation in this study was that infants younger than 7 months of age exhibited the most aggressive and resistant leukemia. Moreover, blast cell morphoimmunology were unique, monocytic, early B-cell, or undifferentiated mixed, reflecting a selection of stem cells in active proliferative hemopoiesis typical in the neonatal period. It is anticipated that further investigations may shed additional insight into the specific mechanism governing the unique drug-resistant stem cell leukemia of early infancy.

124
REFERENCES

CANCER January I 1989

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