A case of acute lymphoblastic leukemia with severe hypodiploidy

A Case of Acute Lymphoblastic Leukemia with Severe Hypodiploidy Shinichi Misawa, Nobuo Oguma, and Joseph R. Testa

ABSTRACT: A case of acute lymphoblastic leukemia with a severe hypodiploid chromosome constitution is reported. The modal chromosome number was 36, and the karyotype of these cells was 3 6 , X , - X , - 2 , - 3 , - 5 , - 7 , - 9 , - 1 2 , - 1 3 , - 1 5 , - 1 6 , - 1 7 , - 2 0 , + 2 1 , + mar,del(1) (p13.1p22.3),inv(3)(q13.3q29). In addition to a haploid set, extra copies of chromosomes #6, #10, #14, #18, and #21 were found, as in most cases with severe hypodiploid karyotypes. A second, near-triploid cell line was also observed. An examination of chromosomal heteromorphisms suggested that the severe hypodiploid clone originated either from a near-triploid cell or from a common precursor cell.

INTRODUCTION Severe h y p o d i p l o i d y (less than 40 chromosomes), i n c l u d i n g near-haploidy, is a rare occurrence in h u m a n leukemia. In several large series of leukemia patients studied with chromosome b a n d i n g methods, severe hypodiploidy was observed i n 3 of 346 cases (0.87%) or 1 of 138 cases (0.74%) with acute lymphoblastic leukemia (ALL) [1, 2] and in 1 of 158 cases (0.63%) with chronic myelogenous leukemia (CML) i n blastic crisis [3]. We know of no report of severe h y p o d i p l o i d y in acute n o n l y m phocytic leukemia, although there have been reports of extreme h y p o d i p l o i d y in two cases of either myeloid or myelomonocytic blastic crisis of CML [4, 5]. The lowest chromosome n u m b e r reported in h u m a n leukemia is 24, seen in one patient with ALL and in another with CML in blastic crisis [6, 7]. Each of these cases showed a haploid set of autosomes and an XY sex chromosome constitution. Other severe hypodiploid cases retained two copies of some autosomes (especially chromosomes #6, #10, #14, #18, and #21) as well as two sex chromosomes i n most instances [8]. In this article we describe a patient with ALL whose bone marrow contained both severe hypodiploid and near-triploid clones. Possible routes of clonal evolution leading to the two cell populations in this case are discussed. CASE HISTORY A 55-year-old white female was admitted to the University of Maryland Cancer Center on November 19, 1981 with a history of shortness of breath for the previous 3 weeks. The patient had a white blood cell count of 3100/p.1, with 50% blasts, a platelet count of 50,000/v~l, and a hematocrit of 35% after transfusion of red cells. From the CytogeneticsSection,Universityof MarylandCancer Center, Baltimore,MD Address requests for reprints to Dr. Joseph R. Testa, Cytogenetics Section, University of Maryland Cancer Center, 655 W. Baltimore Street, 9-031 BRB, Baltimore, MD 21201. Received March 12, 1984; accepted June 25, 1984.

137 © 1985 Elsevier SciencePublishingCo., Inc. 52 VanderbiltAve., New York, NY 10017

Cancer Geneticsand Cytogenetics16, 137-143 (1985) 0105-4008/85/$03.30

13 8

s. Misawa et al. A p p r o x i m a t e l y 80% of the bone marrow cells were leukemic blasts. There were two populations of blasts. A b o u t 80% of these cells were 14-20 wm in diameter, with slightly basophilic cytoplasm and frequent vacuolization. Nuclear clefting was deep and narrow, and one to three distinct nucleoli were noted. The remaining blasts were smaller, 7-10 wm, with scanty cytoplasm, occasional i n d e n t e d nuclei, and few (none to one) nucleoli. No azur granules or Auer rods were observed in either cell type. No cytochemical staining was done because of the limited s p e c i m e n available. Cell surface markers on the cells, w h i c h were separated from b i o p s i e d bone core, were positive for terminal d e o x y n u c l e o t i d y l transferase and c o m m o n ALL antigen by indirect immunofluorescence methods. The diagnosis was acute l y m p h o b l a s t i c leukemia (ALL-L2 according to the FAB classification [9]). The patient was started on treatment with daunorubicin, vincristine, prednisone, and L-asparaginase. Leukemic cells d i s a p p e a r e d from the peripheral blood with this c h e m o t h e r a p y regimen, but the bone marrow still s h o w e d 50% blasts. The chemotherapy protocol was changed to a combination of prednisone, vincristine, L-asparaginase, and methotrexate. A complete remission was achieved on February 10, 1982, w h i c h lasted 4 months. The patient died on September 28, 1982, 10 months from the time of diagnosis.

CYTOGENETIC EXAMINATION Cytogenetic studies were performed before the initiation of chemotherapy. Chromosome preparations were obtained from a bone marrow b i o p s y that was m i n c e d and then cultured for 24 hr. The cells were exposed to c o l c e m i d (0.06 v.g/ml) for 10 min and harvested according to standard methods. A PHA-stimulated p e r i p h e r a l blood sample was also examined in order to establish the constitutional karyotype. Quinacrine-banded chromosomes were identified according to the ISCN (1978) and ISCN (1981) [10, 11], and karyotypes are expressed as r e c o m m e n d e d u n d e r these systems. Twenty-nine cells were photographed. Twenty-three of these cells had 36 chromosomes, and 6 had 61 chromosomes. Twenty-two cells were c o m p l e t e l y analyzed, and the karyotypes are expressed as follows: 3 6 , X , - X , - 2 , - 3 , - 5 , - 7 , - 9 , - 1 2 , - 13, - 15, - 16, - 17, - 20, + 21, + mar (submetacentric,B-size),del(1)(p13.1p22.3), inv(3)(q13.3q29) (Fig. 1) and 6 1 , X , - X , + l p - , + l p - , + 4 , + 6 , + 8 , + 8 , + d e l ( 9 ) (p12p22),+10,+11,+14,+18,+19,+21,+21,+22,+22 [Fig. 2). A n additional 100 metaphase cells were e x a m i n e d u n d e r the microscope to further assess the p l o i d y of mitotic cells. Sixty-six cells showed severe h y p o d i p l o i d y and 34 near-triploidy. Each of the 20 cells from PHA-stimulated peripheral blood showed a normal female karyotype. RESULTS

AND

DISCUSSION

Acute l y m p h o b l a s t i c leukemia with severe h y p o d i p l o i d y m a y represent a unique clinicopathologic entity associated with a poor prognosis as c o m p a r e d to that in other types of non-T, non-B ALL [8, 12-17]. Even though a complete remission was achieved in seven of eight reported cases, i n c l u d i n g our patient, the m e d i a n survival was only 7 months. Overall, patients with severe h y p o d i p l o i d y show a rather consistent c h r o m o s o m e pattern, especially those with ALL (Table 1). In a d d i t i o n to a h a p l o i d set, + 21 has been seen in 11 of 13 reported ALL cases; +18, +10, + 6 , and + 1 4 in 9, 7, 6, and 5 cases, respectively; and + X or + Y in 11. Thus, in our case, most of the extra chromosomes seen in the severe h y p o d i p l o i d clone have also been observed frequently in other ALL cases [i.e., five of the six most c o m m o n additional chromo-

ALL with Hypodiploidy

139

Figure 1 Karyotypeof a Q-banded bone marrow metaphase cell with 36 chromosomes. Arrows indicate breakpoints involved in interstitial deletion of chromosome #1 and inversion of chromosome #3. somes were present in our case). Interestingly, the pattern of additional chromosomes in severe hypodiploid leukemia is remarkably similar to that seen in ALL patients with hyperdiploid clones with modal numbers of 50-60. Thus, the most consistent changes in the latter group are +21, +6, and +18 [1]. Our patient had two populations of leukemic cells. Similarly, in most other patients with severe hypodiploid leukemia, there is cytogenetic and cytologic evidence for the presence of two populations of leukemic cells, one severe hypodiploid (presumably small blasts} and the other hyperdiploid to near-triploid (large blasts). Eleven of 19 previously reported cases of severe hypodiploid leukemia had a variable percentage of cells that contain double the number of chromosomes of the severe hypodiploid line, nine of which showed an exact doubling of each chromosome (Table 1). We examined chromosomal heteromorphisms in normal diploid, near-triploid, and severe hypodiploid cells to help clarify the route of clonal evolution in our case. Chromosome pairs #3, #9, and #13 showed distinguishable heteromorphisms among homologous chromosomes in PHA-stimulated lymphocytes, representative of the patient's constitutional karyotype. There was brilliant fluorescence near the centromere on both arms of one chromosome #3 and on the short arm only in the other homolog. One chromosome #9 had a large negative (nonfluorescent) segment at 9q12, and the other #9 showed a medium-sized negative band. One chromosome #13 had a brilliant short arm, whereas the short arm of the homolog had medium fluorescence intensity {Fig. 3A). In near-triploid bone marrow cells with 61 chromosomes, there were two distinguishable #3 chromosomes that

140

s. Misawa et al.

Figure 2 Karyotype of a Q-banded marrow metaphase cell with 61 chromosomes. Arrows indicate deletions of chromosomes #1 (two copies) and #9. showed the same h e t e r o m o r p h i s m s as those in lymphocytes. Two normal chromosomes # 9 had a large negative band at 9q12, whereas a third, rearranged # 9 ( 9 p - ) had a m e d i u m - s i z e d negative band. One chromosome #13 had a brilliant short arm and the other had a short arm of m e d i u m fluorescence (Fig. 3B). In severe h y p o d i p loid cells, there were only single copies of chromosomes #3, #9, and #13. These cells showed brilliant fluorescence on the short arm of c h r o m o s o m e #3, a large negative band at 9q12, and a brilliant short arm on #13 (Fig. 3C). Furthermore, a marker c h r o m o s o m e and an inversion of c h r o m o s o m e # 3 were present only in severe h y p o d i p l o i d cells. The findings from the studies of chromosomal h e t e r o m o r p h i s m s indicate that the near°triploid cells could not have originated from a doubling of the chromosomes of the h y p o d i p l o i d cells. This is in contrast to m a n y other reported cases in w h i c h a h y p e r d i p l o i d clone existed with an exact doubling of each c h r o m o s o m e present in the near-haploid clone. In such cases, it was considered most likely that the nearh a p l o i d cells represented the stem line of the leukemia and that the h y p e r d i p l o i d cells evolved from these cells [13, 15]. Chromosomal h e t e r o m o r p h i s m studies have been described in only one previous case of ALL with n e a r - h a p l o i d y [13]. In that case, cells with 54 chromosomes b a d two #22 chromosomes with bright fluorescence on their satellite regions, and cells with 27 chromosomes had a single copy of #22 that had a brightly fluorescent satellite region. On the other hand, normal d i p l o i d cells showed heteromorphic patterns on the satellite regions; one was bright and the other pale. Furthermore, there was a marker in cells w i t h 27 chromosomes and two copies of this marker in cells with 54 chromosomes. These findings suggested that the h y p e r d i p l o i d cells arose from the near-haploid cells.

63/M 17/F 17FF 11/F 52/M 3/F 18/M 16/F 7/M 67/M

55/M 58/M

44/M 55/F 61/F 67/M 14/M

28/1:

3 4 5 6 7 8 9 10 11 12

13 14

15 18 17 18 19

20

No. of

(35.5l

35 26 b 28 b 27 b 28 3 5 - 3 6 b'h

36 b'd 24 b

32 b 28 b 26 b 286 36 26 26 26 24 34 c

27 b

27 b

chromosomes

Sex

1.8

XY XX X,-X X, - Y XY XX

X, X XY

XY XX XX XX XY XX XY XX XY XY

X,-X

XX

chromosomes

2.0

1 1 2 1 1 1 2 1 1 1 1 1 2 1 2 1 1 1 1

1

2.1

1 1 1 1 1 1 1 1 1 1 1 2 1 1 2 1 1 1 1

2

1.O

1 1 1 1 1 1 1 1 1 1 1 0 1 1 1 1 1 1 1

3

1.1

1 1 1 1 1 1 1 1 1 1 1 2 2 1 1 1 1 1 1

4

#11

a r e a v e r a g e o f 15 c e l l s c o m p l e t e l y

1 1 2 2 1 1 2 1 1 2 1 2 2 1 2 1 O 1 1

6

2.1

#20,

analyzed.

#22

2.1

1 p+ 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1

7

2.0

1 1 1 1 1 1 2 1 1 1 1 0 2 1 2 2 1 2 1

8

and inv(3).

#21, and #22

9

1.O

2 2 2 2 1 2 2 1 1 1 1 0 2 1 2 1 1 2 1

10

1.1

1 1 1 1 1 1 2 1 1 1 1 2 2 1 1 1 28 1 1

11

1.5

1 1 1 1 1 1 1 1 1 1 1 2 1 1 2f 1 1 2 1

12

1.5

1 2 1 1 2 2 1 1 2 1 1 1 2 1 2 1 2 1 2

14

1.5

1 1 1 1 1 1 2 1 1 1 1 1 1 1 1 1 1 1 2

15

1.0

1 1 1 1 1 1 1 1 1 1 1 2 1 1 1 1 1 1 1

16

1.1

1 1 1 1 1 1 1 1 1 1 1 2 1 1 1 1 1 1 1

17

cells were also observed.

0.7

1 1 1 1 1 1 1 1 1 1 1 2 1 1 1 1 2 1 1

13

but chromosome

# 9 is n o r m a l .

varied from two to none.

or near-haploid

0.9 e

1 1 2 1 1 1 1 1 1 1 1 2 1 Ie 1e Ie 1 1~ 1e

is P h I c h r o m o s o m e ,

t h e r e is a l p -

are possibly t(11;67). One chromosome

#9 and #22.

p r e s e n t . In a d d i t i o n ,

1.1

1 1 1 1 1 1 2 1 1 1 1 1 1 1 1 1 1 1 1

5

of s e v e r e h y p o d i p l o i d

#19,

of c h r o m o s o m e s

present. Chromosomes

chromosomes

1 2 is 1 2 p - .

~Chromosome numbers

8Two chromosomes

~One c h r o m o s o m e

ePhl t r a n s l o c a t i o n b e t w e e n

dOne consistent marker chromosome

~Fwo c o n s i s t e n t m a r k e r c h r o m o s o m e s

~Cells w i t h d o u b l e o r n e a r l y d o u b l e t h e n u m b e r

°Cases 1-13 are ALL and cases 14-20 are CML in blastic crisis.

4/F

12FF

1

2

Age/sex

Case no. °

Chromosome identification

1.9

2 2 2 2 1 2 2 2 1 2 1 1 2 1 2 1 2 1 1

18

1.8

1 1 2 1 1 1 2 1 1 1 1 2 2 1 1 1 1 1 1

19

1.7

1 1 1 1 1 1 1 1 1 1 1 2 1 1 1 1 1 1 1

20

2.D

2 2 2 2 2 2 2 2 2 1 1 2 3 1 2 2 2 2 2

21

Table 1 Chromosome constitution of near-haploid and severe hypodiploid leukemia studied with banding methods

2.3 e

1 1 2 1 1 1 3 1 1 1 1 2 2 Ie Ie Ie 28 1e 2e

22

P e d e r s e n a n d Boesen [5]

Kessous et al. [12] O s h i m u r a et al. [13] Shabtai et al. [14] Kaneko and Sakurai [15] Brodeur et al. [16] Brodeur et al. [16] Sandberg et al. [8] Hoeltge et al. [17] Chaganti [6] Chaganti [6] Chaganti [6] Chaganti [6] Present case Daniel et al. [7] Como and Graze [4 l H a r t l e y and Cook [18] Kessous et ai. [19] /shihara et al. 120] Najafzadeh et al. [21]

References

g

0 ~-~.

>

142

s. Misawa et al.

Figure 3 Chromosome pairs #3, #9, and #13, which show heteromorphisms among homologous chromosomes from a normal diploid cell (lymphocyte) (A) and #3, #9, and #13 from a near-triploid cell (B) and a severe hypodiploid cell (C). Arrows indicate breakpoints of rearranged chromosomes. In our patient, the modal chromosome n u m b e r of the hypodiploid cells is considerably more than half that of the near-triploid cells, but it is possible that the hypodiploid clone was derived from a near-triploid cell. Alternatively, the severe hypodiploid and near-triploid populations could both have originated from a comm o n precursor cell. For example, both of these clones could have been generated from a pseudodiploid cell containing a l p - abnormality, even though we found no evidence of such a pseudodiploid cell in our cytogenetic examination. Although distinguishing rearrangements are u n i q u e ta both the severe h y p o d i p l o i d (inversion 3, B-size marker) and the near-triploid ( 9 p - ) clones, the c o m m o n derivation of the two cell lines is supported by the presence of the l p - in each. This work was supported in part by PHS grant 1P50CA-32107-01 awarded by the NCI, DHHS. J.R.T. is a Special Fellow of the Leukemia Society of America.

REFERENCES 1. The Third International Workshop on Chromosomes in Leukemia (1981): Chromosome abnormalities in acute lymphoblastic leukemia: Structural and numerical changes in 234 cases. Cancer Genet Cytogenet 4:101-110. 2. Williams DL, Tsiatis A, Brodeur GM, Look T, Melvin SL, Bowman WP, Kalwinsky DK, Rivera G, Dahl GV (1982): Prognostic importance of chromosome number in 136 untreated children with acute lymphoblastic leukemia. Blood 60:864-871.

ALL w i t h H y p o d i p l o i d y

143

3. Ishihara T, Sasaki M, Oshimura M, Kamada N, Yamada K, Okada M, Sakurai M, Sugiyama T, Shiraishi Y, Kohno S (1983): A summary of cytogenetic studies on 534 cases of chronic myelocytic leukemia in Japan. Cancer Genet Cytogenet 9:81-92. 4. Como RM, Graze PR (1979): Emergence of a cell line with extreme hypodiploidy in blast crisis of chronic myelocytic leukemia. Blood 5 3 : 7 0 7 - 7 1 1 . 5. Pedersen B, Boesen AM (1983): Extreme hypodiploidy in a case of myelomonocytic crisis of chronic myelogenous leukemia. Cancer Genet Cytogenet 9:101-112. 6. Chaganti RSK: Quoted by Sandberg et al as a personal communication [8]. 7. Daniel A, Francis SE, Stewart LA, Barber S (1978): A near haploid clone: 24,XY, t(9;22)(q34;q11) from a patient in blast crisis of chronic myeloid leukaemia. Scand J Haematol 21:99-103. 8. Sandberg AA, Wake N, Kohno S (1982): Chromosomes and causation of human cancer and leukemia. XLVII. Severe hypodiploidy and chromosome conglomerations in ALL. Cancer Genet Cytogenet 5:293-307. 9. Bennett JM, Catovsky D, Daniel MT, Flandrin G, Galton DAG, Gralnick HR, Sultan C (1976): Proposal for the classification of the acute leukemias. Br J Haematol 33:451-458. 10. ISCN (1978): An international system for human cytogenetic nomenclature (1978). Cytogenet Cell Genet 21:309-404. 11. ISCN (1981]: An international system for human cytogenetic nomenclature. High-resolution banding (1981). Cytogenet Cell Genet 31:1-23. 12. Kessous A, Corberand J, Grozdea J, Colombies P (1975): Clone cellulaire a 27 chromosomes dans une leucemie aigue humaine. Nouv Rev Fr Hematol 15:73-82. 13. Oshimura M, Freeman, AI, Sandberg AA (1977): Chromosomes and causation of human cancer and leukemia. XXIII. Near-haploidy in acute leukemia. Cancer 40:1143-1148. 14. Shabtai F, Lewinski UH, Har-Zahav L, Gafter U, Halbrecht I, Djaldetti M (1979): A hypodiploid clone and its duplicate in acute lymphoblastic leukemia. Am J Clin Pathol 72:1018-1024. 15. Kaneko Y, Sakurai M (1980): Acute lymphocytic leukemia (ALL) with near-haploidy--A unique subgroup of ALL? Cancer Genet Cytogenet 2:13-18. 16. Brodeur GM, Williams DL, Look AT, Bowman WP, Kalwinsky DK (1981): Near-haploid acute lymphoblastic leukemia: A unique subgroup with a poor prognosis? Blood 58:1419. 17. Hoeltge GA, Dyment PG, Slovak ML (1982]: Acute lymphocytic leukemia with microblastosis and near haploidy (26 chromosomes): A case report. Med Pediatr Oncol 10:53-59. 18. Hartley SE, Cook MK (1979): Near-haploidy in a case of chronic myeloid leukemia. Cancer Genet Cytogenet 1:169-176. 19. Kessous A, Colombies P, Pris J, Clement D (1980]: Near-haploid cell line in lymphoid blast crisis of Phi-positive chronic myeloid leukemia. Cancer Res 40:1354-1359. 20. Ishihara T, Minamihisamatsu M, Kohno S (1982): Near-haploid conversion in Phi-positive chronic myelocytic leukemia. Proc Jpn Acad 58(B):165-168. 21. Najafzadeh TM, Dumars GE, Dumars KW, Simpkins H, Katz J (1983): Near-haploid cell line in the blastic crisis of chronic myelogenous leukemia: A possible marker for lymphoid malignancy. Cancer Genet Cytogenet 9:333-339.