Near-Haploid Clones in a Malignant Fibrous Histiocytoma Charlotte Orndal, Nils Mandahl, Birgitta Carl6n, Helena Will6n, Johan Wennerberg, Sverre Heim, and Felix Mitelman

ABSTRACT: Near-haploid solid tumors are very rare. In a storiform-pleomorphic malignant fibrous histiocytoma (MFH) of bone, we found three cell populations: one with a near-haploid, a second with a near-diploid, and a third with a near-tetraploid chromosome number. The near-haploid cells had f e w structural rearrangements: i(12p) Qnd t(13q21q) in one clone, and these two and an additional t(19;?)(p11;?) in another clone. One structurally normal copy of all chromosomes was also present, except that the only chromosome 13 was involved in the t(13q21q). There were also two near-diploid clones, one without the t(19;?) and one with a single copy of this derivative chromosome. This is the first MFH reported to have a near-haploid modal chromosome number, and also the first tumor with i(12p) among bone and soft tissue tumors.

INTRODUCTION

PATIENT AND METHODS

Malignant fibrous histiocytoma (MFH) of soft tissue is the most common sarcoma of adults [1]. MFH can also be a primary bone lesion. Intraosseous MFHs are often found in areas of previous bone infarction and are aggressive tumors with worse prognosis than those of soft-tissue origin [2]. The storiform-pleomorphic subtype is the most common and also the most aggressive among both soft-tissue and intraosseous MFH [1, 3]. Although MFH of bone is primarily found in long-tubular bones, several cases arising from deep structures of the head and neck have been reported. These tumors tend to be locally invasive, irrespective of histologic subtype [4]. Altogether 32 MFHs with karyotypic anomalies have been reported [5-10]. All of these, except one [7], were softtissue tumors, and the majority had massive numerical and structural chromosome rearrangements. The chromosome number was 32-37 in one tumor, but varied from neardiploid to near-hexaploid in the other tumor, with neardiploid and near-triploid numbers as the most common. We present here the cytogenetic findings in a storiformpleomorphic MFH of bone, which had near-haploid, neardiploid, and near-tetraploid clones.

Clinical and Histopathologic Findings A 52-year-old woman with rheumatoid arthritis, but without previous malignant disease, had a painless tumor in the retromolar trigone of the left side of the mandible. Fine needle aspiration cytology revealed a malignant tumor of mesenchymal origin. X-ray examination showed a destruction in the mandible. No regional or distal metastases were found. A hemimandibular resection was performed. The 3 × 3 x 3 cm tumor had large macroscopic necroses. Microscopically the tumor was highly vascularized and had focal areas of hemorrhage. An abundance of atypical mitotic figures and multinucleated giant cells of tumor type were detected in areas of pleomorphism, and in some areas storiform pattern was found (Fig. 1). Immunohistochemistry by the indirect peroxidase method with antibodies against Sd00 protein, keratin (AE1/AE3, Cam 5.2), factor VIII, desmin, and smooth muscle antigen was negative, but cells positive for a-l-chymotrypsin and a-l-antichymotrypsin were found. Ultrastructural examination revealed fibroblast-like cells, histiocytes, and myofibroblasts distributed in a collageneous matrix (Fig. 2). The histopathologic diagnosis was intraosseous MFH of the storiform-plemorphic type, malignancy grade IV (on a four-grade scale). Twelve months after the primary operation, the patient had a subcutaneous metastasis in the ipsilateral side of the neck, which was removed by a wide local excision. There were still no signs of distal metastases. After another 5 months, a metastasis, or maybe a local recurrence, was found below the petrosus bone, close to the external auditory canal, which was also surgically removed. None of the recurrent tumors were cytogenetically analyzed.

From the Departments of Clinical Genetics (C. (3., N. M., S. H., F. M.), Clinical Pathology (B. C., H. W.), and Oto-Rhino-Laryngology (J. W.), University Hospital, Lund, Sweden, and the Department of Medical Genetics (S. H.), Odense University, Odense, Denmark. Address reprint requests to: Charlotte Orndal, Department of Clinical Genetics, University Hospital, S-221 85 Lund, Sweden. Received November 12, 1991; accepted December 16, 1991.

147 © 1992 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York, NY 10010

Cancer Genet Cytogenet 60:147-151 (1992) 0165-4608/92/$05.00

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C. Orndal et al. errant metaphases with a near-haploid, near-diploid, or near-tetraploid chromosome number. Two clones were found among the near-haploid cells: 11 cells with 24,X,+i[12p),t(13q21q),+21 and 10 cells with 25,X, +i(12p),t(13q21q),+t(19;?)(p11;?),+21 (Fig. 3). Ten cells each had the related near-diploid karyotypes 48,XX, + i(12p), + i(12p),t(13q21q),t(13q2 lq), + 21, + 21, and 49,XX, +i(12p), +i(12p), t(13q21q), t(13q21q), +t(19;?)(p11;?) + 21, + 21. The only one of the ten near-tetraploid cells that was possible to karyotype had four copies of i(12p), one t(13q21q), and two t(19;?)(p11;?).

DISCUSSION Figure 1 Histopathologic section of the tumor showing a fibroblast-rich area with a storiform cell pattern (HE; × 100).

Cytogenetics Fresh tumor tissue was minced with scissors and then treated with collagenase II (1,000 U/ml) for 3 hours. The resulting cell suspension was plated on glass chamber slides in RPMI 1640 medium with HEPES buffer supplemented with 17% fetal calf serum, glutamine, and antibiotics [11[. After 4-7, days the cultures were harvested in situ. Wright stain was used for G-banding.

RESULTS Of the 87 cells analyzed, 36 had a normal diploid or tetraploid female chromosome complement. There were 51 ab-

Near-haploid, mostly hyperhaploid, karyotypes have been reported in 29 hematologic neoplastic disorders [references in 12]. The majority were acute lymphatic leukemia and Philadelphia-positive chronic myeloid leukemia (CML). All had only numerical changes, except for the t(9;22) in CML, with one copy of most chromosomes, but two copies of some. Two sex chromosomes were found in almost all cases and, in falling order of frequency, there was disomy for chromosomes 21, 18, 14, 8, and 10. Including the present case, only 11 solid tumors, representing a variety of histologic types, with at least one nearhaploid clone have been reported (Table 1). True nullisomy does not seem to exist; all cases with chromosome losses in relation to the haploid chromosome number had marker chromosomes that may contain material from the chromosome with apparent nullisomy. Most cases also had related clones with higher ploidy level. Numerical changes were more c o m m o n than structural changes. The most frequent

Figure 2 Electron microscopic pictures showing (A) a mixture of fibroblast-like cells and histiocytes (original magnification x 7020), and (B) a myofibroblast surrounded by collagen ( x 8320).

Near-Haploid Clones in Malignant Fibrous Histiocytoma

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13

14

15

:1

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19

20

)

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11

12

16

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Figure 3

Karyogram of a cell with 24,X,+i(12p),t(13q21q),+21. A der(19) and the normal homologue from a mitosis with 25,X, + i(12p),t(13q21q), + t(19;?)(p11;?], + 21 are displayed in the box.

numerical changes (in relation to the haploid chromosome number) were disomy 21, followed by disomy 7, 19, and 20. It might also be of interest that three structural rearrangements t(13q21q), del(3p), and del(7p), have been found in two tumors each. The near-haploid clones of our case had one structurally unaltered copy of all chromosomes, except that the only chromosome 13 was involved in a t(13q21q). Only segments involved in structural rearrangements, i.e., 12p, 21q, and in one of the clones 19p11--~qter, were present in more than one copy. The tumor cells were thus trisomic for 12p, disomic for 21q and, in half of the cells, 19p11-oqter, and monosomic for all other chromosomes. The near-diploid clones were exact duplications of the near-haploid clones with one exception: the extra der(19) was never present in more than one copy. A cytogenetic picture like that observed in the present case may originate through several mechanisms [see, e.g., 13-17], and it is not possible to conclude with certainty through which pathways the different clones evolved. It

seems likely that the near-haploid clones appeared before those with higher ploidy. The chromosome number might have been reduced through sequential nonrandom losses, multipolar mitoses, or a meiosislike cell division. Whether the structural rearrangements took place before, after, or during the chromosome number reduction, is uncertain. The fact that the der(19) was never present in more than one copy, irrespective of ploidy level, suggests that the near-diploid cells originated through fusion of near-haploid cells with and without the marker. However, polyploidization through acytokinetic mitosis or endoreduplication, followed by loss of one copy of the der(19), cannot be ruled out. In contrast to the present case, most grade IV MFH have had complex karyotypes with multiple numerical and structural aberrations [8]. Most of these were soft tissue tumors. The only bone MFH reported (case 4 in 7) was hypodiploid and had multiple structural aberrations, none of which are the same as those of the present case. Chromosome arm 19p has been frequently involved in aberrations

150

C. (3rndal et al.

Table 1

S o l i d t u m o r s w i t h at l e a s t o n e n e a r - h a p l o i d c l o n e a

Diagnosis Pleomorphic malignant fibrous histiocytoma Osteosarcoma Clear cell chondrosarcoma Endometrioid carcinoma of the ovary Adenocarcinoma of the breast Renal cell carcinoma Renal cell carcinoma Giant cell glioblastoma Meningioma Astrocytoma

Karyotype of near-haploid clone(s} b 24,X, + 21, + i(12p),t(13q21q)/25,same, + t(19;?)(p11 ;?)

32,X,+ 1, + 3 , - 5 , + 8 , - 13,+ 14,+ 18,+ 19,+21, + 21, - 22,1p + ,del(3)(p?),del(14)(q?}, + 4mar 30,X,+ 5,+ 7,+ 12,+ 19,+ 20, + 21,+ 22

Clones at other ploidy levels 48,XX, + i(12p), + i(12p),t(13q2 lq), t(13q2 lq}, + 21, + 21/49,XX, + i(12p), + i(12p}, t(13q21q),t(13q21q}, + t(19;?)(p11;?), + 21, + 21/ tetraploid 62-63/>100

58-60,XX,+5,+7,+7,+12,+12,+12,+19,+19,

Reference Present report

[18] [17]

+ 20,+ 20,+ 21,+ 21,+ 22, + 22 27,X, + 2, + 7, + 10,t(3;?)(q27;?), + mar/ 29,same, + 12, + 20/30,same, + 12, + 20, + mar

[19]

32-34,XX,i(2q),del(6)(q?),7q + ,del(7)(p?),cx

[20]

31,X,+ 10,+ 13,+ 15,+ 17,+ 1 8 , - 1 9 , + 2 0 , + 21, + t(1 ;2)(q12;p21), + mar 34,XY, + 4, + 5, + 8, + 11, + 14, + 15, + 16, + 19, +20,+22 26,X, + 1, + 7, + 9q + ,11q + , - 15, + 18

[21]

32,X, + 5, + 8, + 9, + 15, + 19, + 22, + del(3)(p13), + del(13)(q21), + mar 23-27,XY, + 1, + 7,r(12)(p13q24), + t(18;?)(q23;?)

68

[16]

50,XX,+1,+1,7p+,7p+,+18,+18,-20

[15]

Polyploid

[22]

51-54,XY, +X, + Y, + 1, + 1, + 7, + 7,del(1)(q12)/

[23]

51-54,same,r(12 )(p13q24)/51-54,same, + t(18;?)(q23;?)/100-200 Malignant melanoma

24,X,r(3)?, + del(7)(p13), + der(7)t(6;7)(p11;q11), r(12)?,t(13q21q),der(14)t(1;14)(q12;p11), der(15)t(1;15)(q11;p11)

48

[24]

° Only cases studied with banding techniques are included. Cases with a very wide range of the chromosome number have been disregarded. b In tumors with two or more clones, same refers to the aberrations indicated in the first clone.

i n M F H . A t(19;?)(p11;?} h a s b e e n f o u n d i n t w o t u m o r s , a n d 19p + m a r k e r s w i t h b r e a k p o i n t s i n 1 9 p 1 3 h a v e b e e n f o u n d i n n i n e t u m o r s i n a s e r i e s of 23 M F H [8]. A n i(12p) h a s b e e n f o u n d i n 90 s o l i d t u m o r s , m o s t l y s e m i n o m a s a n d t e r a t o m a s , b u t n e v e r b e f o r e i n a n y b o n e or s o f t - t i s s u e t u m o r [12]. This work was supported by grants from the Swedish Cancer Society, the Swedish Work Environment Fund, the JAP Foundation for Medical Research, and the Lund University Medical Faculty.

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