Rings, dicentrics, and telomeric association in histiocytomas

Rings, dicentrics, and telomeric association in histiocytomas

Rings, Dicentrics, and Telomeric Association in Histiocytomas N. Mandahl, S. Heim, K. Arheden, A. Rydholm, H. Willen, and Felix Mitelman ABSTRACT: We...

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Rings, Dicentrics, and Telomeric Association in Histiocytomas N. Mandahl, S. Heim, K. Arheden, A. Rydholm, H. Willen, and Felix Mitelman

ABSTRACT: We report clonal karyotypic abnormalities in six of 12 cytogenetically investigated malignant fibrous histiocytomas. Four of eight tumors of the pleomorphic subtype had complex clonal chromosome aberrations, including ring chromosomes, dicentric chromosomes, and~or telomeric associations. No common characteristic aberration could be distinguished. Two of four myxoid tumors had clonal changes: One had one to two ring chromosomes and an extra chromosome #2; another had a supernumerary ring chromosome as the sole abnormality.

INTRODUCTION

Although malignant fibrous histiocytoma (MFH) is the most c o m m o n soft tissue sarcoma in h u m a n s [1], cytogenetic analysis of only two cases, to the best of our knowledge, has been published until n o w [2, 3]. In this report we present the cytogenetic findings in eight pleomorphic MFH and four tumors of the myxoid subtype. CASE REPORTS

Eleven tumors were soft tissue lesions. One (case 4) engaged both bone and muscles and probably originated in the bone. All analyses were made on untreated primary tumors. Clinical data on all 12 patients are summarized in Table 1. Additional information is presented below for all cases with clonal chromosome aberrations; case 1 has been described in a previous report [2].

Case 2

The patient had metastatic disease at diagnosis. Cytogenetic analysis was of the primary tumor. This was lobulated, well circumscribed, and partly necrotic. It was highly cellular, with nuclear pleomorphism, and high mitotic activity. The diagnosis was pleomorphic MFH.

From the Departmentsof ClinicalGenetics (N. M., S. H., K. A., F. M.), Orthopaedic Surgery (A. R.), and ClinicalPathology (H. W.), UniversityHospital, Lund, Sweden. Address requests for reprints to Dr, N. Mandahl, Department of Clinical Genetics, University Hospital, S-221 85 Lund, Sweden. Received June 18, 1987; accepted August 20, 1987.

23 © 1988 Elsevier SciencePublishingCo., Inc. 52 VanderbiltAve., New York, NY 10017

Cancer Genet Cytogenet30:23-33(1988) 0165-4608/88/$03.50

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N. M a n d a h l et al.

Table 1

S u m m a r y of c l i n i c a l data

Case

Sex/Age

Diagnosis

Grade"

Sit{,'

Depth

Size (cm)

1

M 54

P-MFII

IV

Hip

D

4 (0)

2 3

M 70 M 67

P-MFH P-MFtt

IV IV

Foot Hip

D S

10 × 6 × 6 7 × 6 x 6

4

M 62

P-MFI|

Ill

I.l p p e r a r m

D

17 x 3 × 2

5 6

M 58 M 74

P-MFII P-MFH

IV IV

Trunk Upper arm

I) S

10 × 9 × 6 7 × 5 x 4

6 (0)

7

F 76

P-MFIi

IV

Thigh

D

8

F 71

P-MFH

IV

Thigh

D

7 x 7 x 3

M-MFH M-MFH MFS MFS

III Ill II II

Groin I , o w e r leg Abdominal wall Forearm

D S S S

8 4 5 1

9 l0 11 12

M M M F

60 69 48 81

× 8 × 7 × 3 x 1 × 4 × 4 (0)

IqliMologic grade of maligmmcy, wher~ IV denotes the highest malignancy grade,.

P-MFH, pleolnorphi(: malignant fibrous histio~ yhmm; M-MFH. IIlyxoid malignant fibrous histiocytoma; MFS. myxofibrosarcoma: I), deep: S. superticial; [t)]. (]itltlit!t(~l'.

Figure 1

Case 3: Pleomorphic type of MFH. tt&E × 300

Rings, Dicentrics, and Telomeres in Histiocytoma

25

Case 3 The tumor was largely necrotic. Some parts consisted p r e d o m i n a n t l y ot spindle cells organized in a storiform pattern; other areas were d o m i n a t e d by a p o p u l a t i o n of bizarre giant cells and a large number of atypical mitoses (Fig, 1). The diagnosis was p l e o m o r p h i c MFH. Case 4

The tumor engaged both bone and surrounding soft tissue. It was mainly composed of s p i n d l e cells, often with a characteristic storiform pattern. The number of mitoses and giant cells varied between different sections. Osteoclastic giant cells were common. Lower cellularity and myxoid areas were found in the periphery. The diagnosis was p l e o m o r p h i c MFH. Case 9 The tumor was partly m y x o i d with some large necrotic foci. The solid and the' m y x o i d structures were intermingled (Fig. 2). In the solid parts the cellularity was

Figure 2

Case 9: Myxoid type of MFH. tt&E × 300

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N. Mandahl et al.

high, with a fairly uniform spindle cell population and some giant multinculear tumor cells. Mitotic cells were rare. The diagnosis was myxoid MFH. Case 11

The tumor had a mixed structure, i n c l u d i n g myxoid areas and highly cellular parts with a p r o m i n e n t storiform growth pattern (Fig. 3). The cells in the myxoid parts were spindle shaped or triangular. Spindle cells with large, irregular nuclei dominated the more cellular areas. There was no necrosis and low mitotic activity. The diagnosis was myxofibrosarcoma.

METHODS

Fresh tumor tissue was disaggregated for 2-3 hours with 0.8% collagenase. The cells were cultured in RPMI 1640 m e d i u m with Hepes buffer s u p p l e m e n t e d with17% fetal calf serum, glutamine, and antibiotics. The cultures were harvested after 3-11 days, and slides were aged at room temperature for 1-4 weeks prior to G-banding hy t r y p s i n - G i e m s a or by Wright's staining. Clonal chromosome abnormalities were defined as gain of a particular chromsome or presence of the same structural rearrangement in at least two metaphases, or loss of a particular chromosome in at least three metaphases. RESULTS

Cytogenetic analysis was attempted on 12 tumors. Clonal chromosomal rearrangements were found in six cases: three had both normal karyotypes and metaphases with nonclona[ aberrations, one case had only normal metaphases, and two cases

Figure 3 Case 11: Myxofibrosarcoma. (Left) section with low cellularity and myxoid background; (right) section with storiform cellular pattern and myxoid background. H&E x 210

27

Rings, D i c e n t r i c s , a n d T e l o m e r e s i n H i s t i o c y t o m a

f a i l e d to y i e l d a n y m i t o s e s . T h e c y t o g e n e t i c f i n d i n g s are p r e s e n t e d b e l o w . Full k a r y o t y p e d e s c r i p t i o n s are g i v e n in T a b l e 2.

Case 1

T h i s c a s e h a s b e e n r e p o r t e d p r e v i o u s l y [2]. T h e c h r o m o s o m e n u m b e r w a s h y p e r d i p l o i d , a n d t h e r e w a s a h i g h f r e q u e n c y of t e l o m e r i c a s s o c i a t i o n s ( m e a n n u m b e r , 2.5/ metaphase).

Case 2

Of 28 m e t a p h a s e s a n a l y z e d , e i g h t h a d a b n o r m a l k a r y o t y p e s w i t h a c h r o m o s o m e n u m b e r b e t w e e n 64 a n d 69. A t o t a l of t e n c l o n a l m a r k e r c h r o m o s o m e s , w h o s e origin c o u l d b e c o m p l e t e l y (four) or p a r t l y (six) i d e n t i f i e d , w e r e f o u n d (Fig. 4). C h r o mosomes #1, #8, #10, #12, #15, #16, and #19 were involved. A small marker r i n g c h r o m o s o m e , i n t e r p r e t e d as r(19), w a s p r e s e n t i n a d d i t i o n to t w o n o r m a l # 1 9 in all m e t a p h a s e s . A f e w t e l o m e r i c a s s o c i a t i o n s w e r e o b s e r v e d ( m e a n n u m b e r , 0.4/ metaphase).

Case 3

Of 76 m e t a p h a s e s a n a l y z e d , 70 h a d a b n o r m a l k a r y o t y p e s w i t h c l o n a l a b e r r a t i o n s . Two clones could be distinguished; one hypodiploid and one hypotriploid. Numerical c h a n g e s c o m m o n to b o t h c l o n e s w e r e losses of c h r o m o s o m e s # 6 , # 1 3 , # 1 7 , Table 2

Case

K a r y o t y p e s of m a l i g n a n t f i b r o u s h i s t i o c y t o m a s

Culture time (days)

l"

4

2

11

3

3 and 4

4

4

5 6 7 8 9 10 11 12

4 4 3 3 3

and and and and and 5 3 and 4

5 9 5 5 6 4

Karyotype 50-56,X, + X , - Y, + 4, + 5,t(11;?)(p15;?),- 15, + 18, + 18, + 20, + 20, + 22, + 22, + 1-5 mar (2.5 tas/metaphase) 64-69,XY, + X, + der{1)t(1;?)(p34;?), + 2, + 3, + 4, + 5, + 5, + 6, + 7,8q +, + 8q +, +del(8)(q13),+9,t(10;12)(p15;q14),+11,+12, 15, 15, + der(15)t{15;?)(p?;?), + der(15)t(15;?)(p?;?), + der(15)t(15:?)(p?;?), + der(16) t(16:?)(q24:?), + r(19)(p13q13), + 20, + 2-4 mar (0.4 tas/metaphase) 32-37,X,-Y,del(1)(qll),-2,i(4q), 5 , - 6 , 1 1 , + d i c { l l ; 2 2 ) ( p l l ; p 1 1 ] , - 1 2 , 13, 14, 1 5 , - 1 6 , 17, 1 8 , - 1 9 , 1 9 , - 2 0 , - 2 1 , - 2 2 , + r , + 2 - 3 mar 54-68,X, + X, - Y,del(1)(ql 1), + del(1){ql 1), + 3, + i(4q), + i(4q), 6,+8, + 9 , + d i e (11;22)(p11;p11),-13, 17, 1 8 , - 1 9 , 20,+10-15 mar (2.2 tas/metaphase) 42,XY, - 1, + der(1)t(1 ;10)(p11;ql 1 ), - 6, + der(6)t(6;?)(pl 1 ;?), 7, + der(7)t(7;?) (p22;?),-7,t(8;9)(q22;p24),-10, 10,+der(10)t(10;?)(q11;?), 12,+der(12) t(12;?)(q24;?),- 15, + dic(1;15)(p13;q26), + der(16)t(16;?)(pl 3;?), - 1 7 , - 19, + der(19)t(19;?)(p13;?), 2 0 , - 20,r(21)(p13q22),r(22)(pl 3q13), + 1-2 mar 46,XY 46,XY Failure Failure 47-48,XY, + 2, + 1-2 r 46,XY 47,XY, + r 46,XX

"This case has been re.porled previously (Mandahl el al. 1985).

28

N. M a n d a h l et al.

-i i-i tt 1

2

3

5

4

lZ'-6

7

imtl

14

~" 19

M1

9

10

11

12

16

17

18

I

13

b

_ 8

X

15

.--

't 20

M2

22

21

M3

M4

M5

M6

M7

Z|,

M8

X Y

M9

MIO

Figure 4 Case 2:(a) Representative karyotype demonstrating all ten clonal and three nonclonal marker chromosomes. Arrowheads indicate breakpoints. (b) The ten clonal marker c:hromosomes selected from different metaphasos. MI: der(l)t(1:?)(p34;?); M2: del(8)(q13): M3: 8q+: M4: der(10)t(10:12)Ip15:q14); M5: der(12)t(10:12)(p15;q14): M6 M8: all different der(15)t[15;?)(p?;?): M9: der(16)t(16;?](q24;?): N110: r(19)(p13q13).

#18, #19, #20, and Y. Both also had a large number of telomeric associations involving several different telomeres (mean number, 2.2/metaphase). The following structural changes were found in the h y p o d i p l o i d clone: del(1)(q11), i(4q), die(l 1;22)(pl 1;p11), a small ring chromosome, and 2-3 markers of u n k n o w n origin. The h y p o t r i p l o i d clone bad the same three identifiable markers, del(1) and the i(4q) in duplicate, no ring, and as many as 10-15 unidentifiable markers (Fig. 5). These and other d u p l i c a t i o n s of markers indicate that the h y p o t r i p l o i d clone emanated from the b y p o d i p l o i d by polyploidization, w h e r e u p o n cytogenetic evolution proceeded i n d e p e n d e n t l y in the two clones. Case 4 All 27 metaphases analyzed had abnormal karyotypes. The chromosome number was h y p o d i p l o i d with remarkably small variation between cells. No telomeric association was observed. A large, stable dicentric chromosome, made up of the entire c h r o m o s o m e #15, and the long arm and part of the short arm of chromosome #1, was present in all metaphases (Fig. 6). Two small ring chromosomes, interpreted as r(21) and r(22), were found in ten of 12 fully k a r y o t y p e d metaphases. In eight different translocations, some c o m p l e t e l y and others only partly characterized, nine of ten identified breakpoints were located in telomeric or centromeric bands.

29

Rings, D i c e n t r i c s , a n d T e l o m e r e s in H i s t i o c y t o m a

4 j-C¢ 1

2

|t[ 4

3

l M3

5

M5

M4

_;tP

j

6

7

8

9

10

11

12

i

a

13

14

19

20

15

'l' b

M1

M2

16

17

21

22

i

A

18

X Y

4

M3

M4

MS

M6

Figure 5 Case 3:(a) Karyotype from the hypotriploid clone showing five (M1 M5) of the six clonal and several nonclonal marker chromosomes. Telomeric associations involve the telomeres of 6q and 4p, l l q and a nonclonal marker, 15p and another nonclonal marker. The long chain of chromosomes include all of chromosomes #12 and #17, material of unknown origin and part of #3 (p21--,qter), associated or fused to each other. Arrowheads indicate breakpoints. (b) The six clonal marker chromosomes selected from both the hypodiploid and the hypotriploid clone. MI: del(1)(q11); M2: i(4q); M3: dic('ll;22)(p11;p1]); M4-M6: unidentified clonal markers. M6 is a small ring chromosome.

Case 5

T h i r t e e n m e t a p h a s e s w e r e a n a l y z e d . A b n o r m a l k a r y o t y p e s w e r e f o u n d ill s e v e n m i toses: six h a d r a n d o m loss or g a i n a n d o n e h a d s e v e r a l s t r u c t u r a l l y r e a r r a n g e d c h r o mosomes. Thus, no clonal aberration was detected.

Cases 6-8

In case 6 o n l y t w o n o r m a l m e t a p h a s e s w e r e f o u n d , w h e r e a s , in cases 7 a n d 8 11o metaphases were found.

Case 9

O n l y t e n m e t a p h a s e s c o u l d b e at least p a r t l y a n a l y z e d ; five of t h e m h a d 1 or 2 r i n g c h r o m o s o m e s . Of f o u r k a r y o t y p e d m e t a p h a s e s t w o w e r e n o r m a l , one was 46,X,-Y,+2, 11,-12,-12,-22,+2r,+2mar (Fig. 7a), t h e f o u r t h 46,X, Y , + 2 , - 1 1 , - 1 2 , + r , + m a r (this m a r k e r w a s p o s s i b l y a d i s t o r t e d c h r o m o s o m e # 1 1 or # 1 2 ) . T h e s e f i n d i n g s are too d i s p a r a t e to d e f i n e a c l o n e w i t h c e r t a i n t y ,

30

N. Mandahl et al.

1

2

3

6

7

8

9

10

4

5

11

12

--:.--1J-13

i 15 I I

14

16

17

Ak 21

4~ 22

18

If_., 19

20

))i=¢, b

M1

M2

M3

M4

M5

~t xY

| ,i = ; , -

,

M6

M7

I1

M8

M9

MIO

Mll

M12

Figure 6 Case 4:(a) Karyotype including all 12 clonal marker chromosomes. Arrowheads indicate breakpoints. (b) The 12 clonal marker chromosomes selected from different metaphases. MI: der(1)t(1;10)(p11;q11); M2: der(6)t(6;?)(pll;?); M3: der(7)t(7;?)(p22;?); M4: der(8)t(8;9)(q22;p24); M5: der(9)t(8;9)(q22;p24); M6: der(10)t(10;?)(q11;?); M7: der(12) t(12;?)(q24;?); M8: dic(1;15)(p13;q26); M9: der(16)t(16;?)(p13;?); M10: der(19)t(19;?)(p13;?); Ml1: r(21)(p13q22); M12: r(22)(p13q13). but tentatively we have accepted gain of c h r o m o s o m e #2 and of one or two ring ch r o m o s o m e s as clonal changes.

Case 10 Of 25 metaphases analyzed four were abnormal. None of the changes were clonal.

Case 11 A total of 26 metaphases were analyzed. In addition to metaphases with normal male karyotype and h y p o d i p l o i d mitoses with r a n d o m losses, a fairly large ring c h r o m o s o m e was found in ten cells, four of w h i c h could be karyotyped. These were 47,XY, + r (two cells), 47,Y, - X, + 13, + r and 47,XY, + 8, - 15, + r (Fig. 7b). The origin of the ring c h r o m o s o m e could not be determined.

Case 12 A total of 38 metaphases were karyotyped. A normal female c h r o m o s o m e complement was found in 21 cells, seven had r a n d o m losses of a variety of autosomes, and ten had various structural aberrations. None of these abnormalities were clonal. The

31

W

a

jo

| 0

b

¸¸¸¸2

J' ~

b Figure 7

[a) Case 9: Metaphase demonstrating the ring chromosomes (arrows) and the small markers (arrowheads). (b) Case 11: Metaphase showing the ring chromosome.

32

N. M a n d a h l et al. only c h r o m o s o m e involved more than once was #17, as 1 7 p + , del{17)(p11) in three different metaphases.

17q+,

and

DISCUSSION

We have analyzed 12 tumors and found clonal c h r o m o s o m e aberrations in four cases of p l e o m o r p h i c MFH and in two m y x o i d tumors, diagnosed as m y x o i d MFH and myxofibrosarcoma. The m y x o i d types of MFH are more often of lower histologic malignancy grade and have better prognosis than p l e o m o r p h i c MFH [1, 4]. The p l e o m o r p h i c tumors had m u c h more c o m p l e x derangements than the tumors with m y x o i d histology, indicating that the n u m b e r and types of chromosomal changes m a y correlate with prognosis. Although a large n u m b e r of clonal structurally rearranged chromosomes were found, no c o m m o n aberration could be identified, nor could any specific numerical aberration be distinguished. The only similarity was restricted to chromosomal behavior, i.e., the presence of ring chromosomes (five of six tumors), dicentric chromosomes (two of six), and/or telomeric associations (three of six). It should be mentioned that telomeric associations were found in three of four p l e o m o r p h i c tumors, but in none of the myxoid. It may be argued that these three rather u n c o m m o n chromosomal p h e n o m e n a are pathogenetically related, and that they all d e p e n d on telomeric association. Intrachromosomal association of telomeres may p r e d i s p o s e to ring chromosome formation; the two forms w o u l d be indistinguishable and w o u l d both be interpreted as rings. A few nonclonal ring c h r o m o s o m e s were observed in cases 1, 2, and 3, i.e., in three of four aberrant p l e o m o r p h i c MFH. On the other hand, interchromosomal association, between telomeres or a telomere and a broken interstitial end, may result in a dicentric chromosome. Observations of telomeric associations in neoplastic cells, their possible origin and significance have only recently been reported and discussed [2, 5, 6]. Also, the related p h e n o m e n o n of association between the telomere of an intact normal c h r o m o s o m e and the centromere of a deleted chromosome has been described in two n e o p l a s m s [5, 7]. The significance of rings, dicentrics, and telomeric associations for tumor initiation or progression remains totally obscure. In particular, this goes for the pleomorphic MFH, in view of their very c o m p l e x rearrangements. It seems more tempting to suggest an important role for the ring chromosomes, being one of two and the only aberration, respectively, in the two myxoid tumors (cases 9 and 11). Only 2% of tumors with cytogenetic abnormalities detected with banding techniques have ring chromosomes [8J. However, this figure is calculated from data mainly based on hematologic malignancies [9], and the situation might be different in soft tissue tumors. Ring c h r o m o s o m e s have for example been identified as characteristic changes in lipoma and well differentiated liposarcoma [8, 10]. It is of interest in this context that in some of the lipomas with ring chromosomes, several telomeric associations were observed (Mandahl, u n p u b l i s h e d observations), though not at such high frequencies as in the p l e o m o r p h i c MFH. If at least some of the ring markers were important in tumorigenesis, this could be achieved through several different mechanisms. The ring of the myxofibrosarcoma (case 11) was an extra chromosome. Consequently, the cells must have been trisomic for one or more c h r o m o s o m e segments. The closure of a linear chromosome to a ring brings genes that are w i d e l y separated on the DNA molecule into close physical proximity, thereby possibly changing the ordinary pattern of gene activity. This w o u l d not differ essentially from other structural rearrangements involving position effects. Duplications or deletions generated through a breakagef u s i o n - b r i d g e cycle [11] may introduce further genomic imbalance.

Rings, D i c e n t r i c s , a n d T e l o m e r e s i n H i s t i o c y t o m a

33

Supported by grants from the Swedish Cancer Society and the JAP Foundation for Medical Research.

REFERENCES 1. Weiss SW (1982): Malignant fibrous histiocytoma. A reaffirmation. Am ] Surg Path 6:773 784. 2. Mandahl N, Heim S, Kristoffersson U, Mitelman F, ROOser B, Rydholm A, Will6n H (1985): Telomeric association in a malignant fibrous histiocytoma. Hum Genet 71:321-324. 3. Turc-Carel C, Dal Cin P, Limon J, Rao U, Li F, Corson JM, Zimmerman R, Parry DM, Cowan JM, Sandberg AA (1987): Involvement of chromosome X in primary cytogenetic change in h u m a n neoplasia: Non-random translocation in synovial sarcoma. Proc Natl Acad Sci USA 84:1981 1985. 4. Merck C, Angervall L, Kindblom LG, Oden A (1983): Myxofibrosarcoma, a malignant soft tissue tumor of fibroblastic-histiocytic origin--A clinical, pathologic and prognostic study of 110 cases using multivariate analysis. Acta Path Microbiol Immunol Scand Sect A 91: suppl No 282. 5. Fitzgerald PH, Morris CM (1984): Telomeric association of chroulosomes in B-cell lymphoid leukemia. Hum Genet 67:385--390. 6. Morgan R, Jarzabek V, Jaffe JP, Hecht BK, Hecht F, Sandberg AA (1986): Telomeric fusion in pre-T-cell acute /ymphoblastic luekemia. Hum Genet 73:260-263. 7. Raimondi SC, Ragsdale ST, Behm F, Rivera G, Williams DL (1987): Multiple telomeric associations of a trisomic whole q arm of chromosome 1 in a child with acute lyinphoblastic leukemia. Cancer Genet Cytogenet 24:87-93. 8. Heim S, Mandahl N, Kristoffersson U, Mitelman F, R66sser B, Rydholm A, Will6n H (1987): Marker ring c h r o m o s o m e - - A new cytogenetic abnormality characterizing lipogenic tumors? Cancer Genet Cytogenet 24:319--326. 9. Mitelman F (1985): Catalog of Chromosome Aberrations in Cancer, 2nd gd. Alan R. Liss, NY. 10. Becher R, Wake N, Gibas Z, Ochi tt, Sandberg AA (1984): Chromosome changes in soft tissue sarcomas. J Natl Cancer inst 72:823 831. 11. McClintock B (1938): The production of homozygous deficient tissues with mutant characteristics by means of the aberrant mitotic behavior of ring-shaped chromosomes. Genetics 23:315 376.