Chromosome Abnormalities in Liposarcomas Yuko Ohjimi, Hiroshi Iwasaki, Yasuhiko Kaneko, Masako Ishiguro, Akiko Ohgami, Chikako Fujita, Norio Shinohara, Kenzo Yoshitake, and Masahiro Kikuchi
ABSTRACT: We performed a cytogenetic study of short-term cultures from fresh surgical specimens obtained from four patients with liposarcoma. Myxoid liposarcomas (cases 1-3) were associated with a specific translocation between chromosomes 12 and 16. Trisomy 8, a nonrandom secondary aberration in myxoid liposarcoma, was observed in the third case as the only additional change. Round cell liposarcoma (case 4) showed complex chromosomal aberrations affecting chromosomes 1, 2, 5, 6, 7, 13, 14, 17, 19, and 22. Neither band 12q13 nor 16pl I was visibly rearranged. Three subgroups of liposarcomas are proposed. The first group is characterized by t(12;16)(q13;p11), the second group by ring chromosomes, telomeric associations, and giant markers, and the last by complex numerical and structural aberrations.
INTRODUCTION Differential diagnosis in soft tissue tumors is often very difficult for both clinicians and pathologists. Special stains, immunohistochemistry, and electron microscopy have been introduced as useful tools for pathologic studies, but poorly differentiated sarcomas remain the most difficult tumors to diagnose [1]. Cytogenetic study is now recognized as a useful technique for diagnosis and estimation of prognosis for blood malignancies. Recently, cytogenetic studies have also proved useful for analysis of solid tumors [2, 3]. Apparently diagnostic chromosome rearrangements have been described in several varieties of malignant soft tissue tumors. These rearrangements include t(X;18)(p11.2;q11.2) in synovial sarcoma [4], t(11;22)(q24;q11.2-12) in Ewing's sarcoma [5] and peripheral primitive neuroectodermal tumors [6], t(2;13)(q35-37;q14) in alveolar rhabdomyosarcoma [7], and t(12;16)(q13;p11) in myxoid liposarcoma [8]. We report the results of chromosome analysis in three cases of myxoid liposarcoma and one case of round cell liposarcoma.
MATERIALS AND METHODS Clinical Data All patients were Japanese, and clinical data of patients are summarized in the Table 1. Case 1 was detailed in another article [9]. From the Department of Pathology (Y. 0., H. I., M. I., A. 0., C. F., K. Y., M. K.I, Fukuoka University School of Medicine, Fukuoka, Laboratory Medicine (Y. K.), Saitama CancerCenter, Saitama, and Department of Orthopedics (N. S.), National Fukuoka Central Hospital, Fukuoka, Japan. Address reprint requests to: Yuko Ohjimi, M.D., Department of Pathology, Fakuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-01, Japan. Received February 21, 1992; accepted May 14, 1992.
Surgical Specimens Tumor tissues were divided immediately into four parts and processed for light and electron microscopy, frozen sectioning, and short-term culturing. Light Microscopy The tumor tissues were fixed in 10% formalin or B-5 fixative. Paraffin sections prepared from these samples were stained with H&E and alcian blue with or without prior hyaluronidase digestion. To detect fat in the tumor cells, cryostat sections were prepared from the frozen tumor tissue and stained with oil red-O. Electron Microscopy The fresh surgical specimens were fixed in 3.0% glutaraldehyde in 0.1 M cacodylate buffer, postfixed in 1.0% osmium tetroxide, dehydrated in ethyl alcohol, and embedded in Epok 812 (Nagase & Co. Osaka, Japan). The sections were cut with a Reichert ultramicrotome, stained with uranylacetate and lead citrate, and examined with a JEM 100 CX-II electron microscope (Jeol, Tokyo, Japan). Cytogenetic Investigation Minced fragments of the tumor tissue were suspended in 200 U/ml type II collagenase (Worthington Biochemical Corporation, Freehold, NJ) for 16 hours [10]. After digestion, the tumor cells were washed and resuspended in the culture medium in glass flasks (TD40, Ikemoto Scientific Technology, Tokyo, Japan), and then incubated at 37°C in a humidified atmosphere containing 5% CO 2 and 5% 02. The culture medium consisted of ES medium "Nissui" (Nissui Pharmaceutical, Tokyo, Japan), supplemented with 10-17% fetal calf serum (HyClone Laboratories, Logan, UT) and 0.3% glutamine (Nissui Pharmaceutical Co., Tokyo, Japan). Colcemid (Life Technologies, Gaithersburg, MD) 111
© 1992 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York, NY 10010
Cancer Genet Cytogenet 64:111-117 (1992) 0165-4608/92/$05.00
112
Table 1
Y. Ohjimi et al.
Clinical and cytogenetic data of liposarcoma patients
Case no.
Primary Age/ tumor sex site
Histologic type
RT or Source CT
Tumor size (cm)
Prognosis (mo)
10 × 10
No. of
Culture metaphases (days) analyzed
1.SFT8925 57/F
R thigh
Myxoid LS
P
No
18, AWD
3
2.SFT9019 39/F 3.SFT9034 35/M
L thigh L thigh
Myxoid LS Myxoid LS
P R
No 5 x 6 11, AWD No 2 × 1.5 × 1 6, AWD
2 5
4.SFT9002 40/M
R thigh
Round cell LS
P
No
2
10 x 10
17, AWD
5 2 33 10 1 8 12
Karyotype 46,XX,t(12;16)(q13;p11) 46,XX 46,XX,t(12;16)(q13;p11) 47,XY, + 8,t(12;16)(q13;p11) 46,XY,t(12;16)(q13;p11) 46,XY 56-62,XY, + 1, + 2, + 5, + 6, + 7 , - 13,- 13,- 14, der(17)t(17;?)(q25;?), + der(17),+ 19,- 22, +9-15 mar
Abbreviations: LS, liposarcoma;P, primary tumor; R, recurrent tumor; RT, radiationtherapy; CT, chemotherapy; AWD.alive without disease.
was added in the last 2 hours at a final concentration of 0.02 /xg/ml. Cells were harvested by trypsinization. After hypotonic shock in 0.075 M KC1 solution for 2 0 - 3 0 minutes, cells were fixed gradually by addition of Carnoy's solution. Cells were spread on moisture slide glasses and dried at room temperature for 2 days. Trypsin-Giemsa b a n d i n g stain was performed. The karyotypes were described according to the short system of the International System for H u m a n Cytogenetic Nomenclature [11].
RESULTS Light Microscopy In case 1, H&E stain showed proliferation of round and polygonal cells with atypical hyperchromatic nuclei. Hypercellular areas with a few mitotic figures were observed. The tumor cells appeared immature, showing no obvious lipoblastic differentiation. Fine networks of capillary blood vessels forming an alveolar pattern were evident in the stroma (Fig. 1A and B). Alcian blue stain demonstrated an a b u n d a n t mucous substance in the intercellular matrix; the substance was digested by hyaluronidase. Oil red-O stain showed only small fat droplets in the cytoplasm of the original tumor. Cases 2 and 3 showed typical histologic features of myxold liposarcoma, with proliferating lipoblasts, delicate plexiform capillary patterns, and an a b u n d a n t myxoid matrix (Fig. 1C and D). Oil red-O stain showed a b u n d a n t and large fat droplets in the cytoplasm. In case 4, round tumor cells with pale or slightly eosinophilic and somewhat vacuolated cytoplasm were arranged in sheetlike pattern with a fine vascular network. Many mitotic figures were apparent among the tumor cells. Oil red-O stain on frozen sections showed small fat droplets in
tumor cells. The histologic feature was indicative of poorly differentiated found cell liposarcoma (Fig. 2).
Electron Microscopy In case 1, the tumor cells generally had few and poorly developed organelles, although some cells contained dilated rough endoplasmic reticulum. The cells possessed a few small n o n m e m b r a n e - b o u n d lipid droplets (Fig. 1E). In cases 2 and 3, the electron microscopic findings were almost the same. Many tumor cells had one or more nonm e m b r a n e - b o u n d lipid droplets of variable size and density and little or no endoplasmic reticulum. The nuclei were large and deeply indented and located at the periphery of the cell (Fig. 1F). In case 4, some tumor cells had lipid droplets, large nuclei with p r o m i n e n t nucleoli, and sparse organelles. There were no desmosomes or other cellular attachments.
Cytogenetic Analysis Results of cytogenetic analysis are summarized in Table 1. Case 1. Of the seven G-banded metaphases analyzed, five
showed the karyotype 46,XX,t(12;16)(q13;p11), with the translocation as the only chromosome change (Fig. 3, left). One cell showed an apparently normal karyotype, 46,XX, and the other showed a karyotype of 4 5 , X X , - 1 5 , which was considered a random missing chromosome. Case 2. Thirty-three analyzable metaphases were obtained.
All showed the clonal (q13;p11) (Fig. 3, center).
karyotype
46,XX,t(12;16)
Case 3. Nineteen metaphases were adequate for cytoge-
netic analysis. Ten of them showed 47,XY,+8,t(12;16)
Figure 1 (A) Delicate capillary network and abundant my×old matrix (case 1, H&E, original magnification x 130). (B) Proliferation of round to polygonal tumor cells with hyperchromatic nuclei (case 2, H&E, original magnification x 515). (C) Delicate plexiform capillary patterns and abundant myxoid matrix are evident (case 3, H&E, original magnification x 130). (D) Proliferating lipoblasts and delicate capillaries (case 3, H&E, original magnification x 515). (E) Electron micrograph of case 1 showing dilated rough endoplasmic reticulum and a few fat droplets (original magnification × 8,550). (F) Electron micrograph of case 3 showing tumor cell with nonmembrane-bound lipid droplets and little endoplasmic reticulum (original magnification x 9,150).
114
A
B Figure 2
Histologic pictures of round cell liposarcoma (case 4). (A) Fine vascular networks and proliferation of round cells (H&E, original magnification × 130). (B) Round tumor cells with pale or slightly eosinophilic cytoplasm (H&E, original magnification x 515).
(q13;p11), one s h o w e d 46,XY,t(12;16)(q13p11), and the other eight showed an a p p a r e n t l y normal karyotype, 46,XY (Fig. 3, right). Case 4. Twelve metaphases were karyotyped. There were many numerical and structural aberrations. Although no two metaphases showed exactly the same karyotypes, many c o m m o n aberrations were noted. A representative karyotype was 5 6 - 6 2 , X Y , + 1 , + 2 , + 5 , + 6 , + 7 , - 1 3 , - 1 3 , - 1 4 , der(17)t(17;?)(q25;?), + tier(17), + 1 9 , - 22, + 9 - 1 5 m a r (Fig. 4).
DISCUSSION Soft tissue (mesenchymal) n e o p l a s m s are rare, accounting for less than 1% of h u m a n malignancy. Unless histopathologists work in large cancer centers, they see few of these tumors. Moreover, undifferentiated or poorly differentiated soft tissue tumors often lack any specific cellular feature.
Y. Ohjimi et al. Recently, cytogenetic analysis of soft tissue tumors has become a valuable tool in diagnosing and defining these tumors and even in providing clues to their histogenesis [2, 3]. Some of these tumors are now k n o w n to have specific c h r o m o s o m e abnormalities. In 1986, Turc-Carel et al. [8] first described a t(12;16)(q13;p11) as a characteristic aberration of m y x o i d liposarcoma, w h i c h is n o w considered one of the tumorspecific c h r o m o s o m e aberrations. In our case 1, pathologic findings such as fine capillary networks, r o u n d to polygonal tumor cells with atypical h y p e r c h r o m a t i c nuclei, and myxold stroma were suggestive of m y x o i d liposarcoma. Ultrastructural examination and oil red-O fat stain failed to demonstrate any evidence of lipoblastic differentiation in the tumor cells, however. Chromosome analysis of the tumor cells in case 1 showed a reciprocal t(12;16)(q13;p11). The existence of t(12;16) confirmed the diagnosis of m y x o i d liposarcoma in this case. Cases 2 and 3 showed typical histologic features of myxoid l i p o s a r c o m a and t(12;16)(q13;p11). Case 3 exhibited an additional numerical aberration: trisomy 8. In the available literature, 28 cases of cytogenetically analyzed m y x o i d and related liposarcomas have been reported [2, 8, 12-22]. We report three additional cases of m y x o i d liposarcoma, including two primary tumors and one recurrent tumor, i.e., 32 tumors in 31 patients (Table 2), including 14 primary, 12 recurrent, and 5 metastatic tumors. The t(12;16)(q13;p11) was apparent in 20 (61%) of the tumors. The same translocation was evident in 12 (85%) of the primary tumors. Aberrations with breakpoints in 12q13 and 1 6 p l l were noted in 26 (81%) and 24 (75%) of the tumors. Although some of the m y x o i d liposarcomas did not show both breakpoints, molecularilevel changes cannot be ruled out for both bands [18]. Eneroth et al. [18] and Streekantaiah et al. [12] reported that trisomy 8 existed as a n o n r a n d o m secondary change in m y x o i d liposarcoma. Mugneret et al. [23] reported that trisomy 8 was observed in 44% of Ewing's sarcomas as well. Trisomy 8 is a c o m m o n secondary numerical abnormality in both chronic and acute m y e l o i d leukemias and indicates a poor prognosis [24]. Case 4 was a r o u n d cell liposarcoma with complex numerical and structural aberrations. Numerical aberrations were evident in chromosomes 1, 2, 5, 6, 7, 13, 14, 17, 19, and 22, with 9 - 1 5 unidentified marker chromosomes and one or two markers derived from chromosome 17. That no two metaphases showed exactly the same karyotype suggests cytogenetical instability in this tumor. Two other cases of cytogenetically analyzed round cell liposarcoma exist [17]; both exhibit c o m p l e x chromosomal aberrations. All three r o u n d cell liposarcomas affected chromosomes 5 and 7 and two affected chromosomes 2, 6, 10, 14, 17, but none of them showed t(12;16)(q13;p11). Therefore, round cell liposarcomas a p p a r e n t l y are cytogenetically different from m y x o i d liposarcomas. Round cell liposarcomas are closely related to myxoid liposarcoma, and most have histologic transitions toward the m y x o i d type. In some instances, the initial tumor is of the m y x o i d type and the recurrent or metastatic tumor is of the round cell type [25]. Four cases of " m i x e d - t y p e "
Myxoid and Round Cell Liposarcomas
115
/
/
1
/
/
/
Figure 3 Partial karyotypes of case 1 (left), case 2 (center), and case 3 (right) showing t(12;16)(q13;p11).
liposarcoma, which probably means the transitional form of liposarcoma, have been reported [2, 18, 21]. Three of them showed t(12;16), and the other showed a complex karyotype without the specific t(12;16). The investigators questioned any cytogenetical relation existed between myxoid, mixed, and round cell liposarcomas. There are three possible answers to this question; First, although histologic transition is observed between myxoid and round cell liposarcomas, there are no cytogenetic transitions. Therefore, myxoid and "mixed" liposarcomas belong to the same group characterized by t(12;16), and round cell liposarcomas belong to another cytogenetic group charac-
terized by complex karyotypes. The second possibility is that round cell liposarcomas have truly mixed abnormalities, including t(12;16)(q13;p11), and that t(12;16)(q13;p11) is masked by complex aberrations. Third, myxoid areas of mixed type have t(12;16)(q13;p11), and round cell areas of mixed type have complex aberrations. We agree with the last answer, because three different clones were found in different parts of one myxoid liposarcoma [20]. We propose the existence of at least three cytogenetic groups of liposarcoma. The first group is characterized by a special translocation, i.e., t(12;p16)(q13;p11). This group has the histologic features of myxoid liposarcoma, and the
Figure 4. Karyotype of case 4 showing complex aberrations: 60,XY,+1,+2,+5,+6,+7,+8,-13,-13,-14, - 17, - der(17)t(17;?)(q25;?) × 2, + der(17;?)(q;?),+ 19, - 22, - 22, + 12mar (arrows point to abnormal chromosomes).
"11 I
!1i 8
7
6
4
5
10
11
12
16
i7
18
3
2
il 9
f
If 14
13
15
f 19 ¸
fl
If 2O
21
22
X
Y
116
Y. Ohjimi et al.
Table 2
Myxoid and related lipasarcomas
Histologic Ref type
Source
RT or CT R
[12] [12]
My My
R P
{121 [12]
My My
R R
[13] [81 [8] [8] [8]
My My My My My
M P P P R
[14]
My
R
[141 [15] [16] [17]
My My My My
R M M R
[171
My My
a)M b)M
[18]
My
P
[18]
My
P
[19] [20]
My My
R PA P B1 P B2
118] [181
Mix Mix Mix Mix
[2] [21]
P B3 P P P
Primary tumor site
Karyotype
R thigh R groin
46,XX,t(12;16)(q13;p11} 46,XY,t(12;16)(q13;p11) 47,XY, + 8,t(12;16)(q13;p11/ R R pelvis 48,XX, + 8,t(11;17)(q13;q21),t(12;16](q13;p11},+ 19 R and C L thigh 46,XY 46,XY,t(12;16)(q13;p11) 46,XY 48,XY,- 9, + 10, - 11, + der(11)t(11;?)(p15;?),t(12;16)(q13;p11),- 17, + 1-3mar R or C R thigh der(12}, 10% of analyzed cells R thigh 46,XX,t(3;15)(p23;q15),t(12;16)(q13;p11) R leg 46,XY,del(6)(q21),t(12;16)(q13;p111 Retroperitoneum 46,XX,t(12;16)[q13;p11) C R thigh 46,XY, + 1,der(1)t(1;12;16)(p11;q13;p11),- 16, der(12)t(1;12;16)(p11;q13;p11) Retroperitoneum del(1)(q11),del(2)(p11),del(3)(p21),t(12;19)(q13;p or q13)[loss der(12)], 6q+,11p+,12p+ Leg 46,XY/46,XY,t(12;16)(q13;p11) ? + der(1)t(1;?)(p11 or p13;?), + 8,t(12;16)(q13;p11),- 16 ? t(12;16;16}(pllq21;p11;q12) Thigh 47,XY,t(1;12)(q42;q13),t(1;16)(q22;p11), + 8 47,XY,+ 8 46,XY,t(11;12}(p15;q13) Thigh 47,XY,- 2,del(6}(q11),t(11;12)(p15;q13),der(15)t(15;?)(q24;?), der(21)t(1;21)(q12;q22), + marl(del(2)?), + mar2 No Thigh 46,XY,der(12)t(12;16)(q13;p11),der(16)t{12;16)(q22;p11) 46,XY No Thigh 46,XX,t(12;16](q13;p11) 46,XX,i(7q),t(12;16)(q13;p11) R and C Retroperitoneum 46,XX,t(12;16)(q13;p11), near tetraploid R thigh 46,XY,t(12;16)(q13;p11) 46,XY,t(12;16)(q13;p11) 46,XY,t(12;16)(q13;p11) 46,XY, + der(1)t(1;16)(p11;p11),t(12;16)(q13;p11),- 16 46,XY 46,XY, + der(1)t(1;12)(p11;q13),der(12)t(12;16)(q13;p11),- 16 47,XY, + 8,t(12;16)(q13.3;p11.2) No Thigh No Thigh 46,XY,t(12;16)(q13.3;p11.2) Thigh Absence of t(12;16); complex karyotype R leg 46,XY,t{12;16)(q13;p11) ?
[121
My+PD
R
]22] My+WD
P
]14] WD+My [14] WD+ My ]14] WD+My
P R R
L thigh
47,XY, + 8,t(12;16)(q13;p11) 94,XXYY, + 8, + 8,t(12;16)(q13;p11),t(12;16)(q13;p11) endoredup|ication with t(12;16) Rthigh 46,XY,t(12;16)(q13;p11} 46,XY Retroperitoneum 11q + ,ring,tas,giant markers Retroperitoneum Near triploid, UMs Retroperitoneum Ring,tas,giant markers
Abbreviations: My, myxoid;Mix, mixed; PD, poorly differentiated;WD, well differentiated;tas, telomeric associations;UMs, unidentifiedmarkers; other abbreviationsas in Table 1. tumor frequently occurs in the thigh. In 1986, Turc-Carel et al. [8] suggested that tumors with this translocation might have a better prognosis. The second group consists of liposarcomas with ring chromosomes, telomeric associations, and giant markers [14]. They have a tendency to occur in the retroperitoneum and histopathologically exhibit the features of n o n m y x o i d (differentiated or pleomorphic) lipo* sarcomas. The third group is characterized by complex numerical and structural aberrations. The tumors often
have the histologic feature of r o u n d cell liposarcoma, characterized by r o u n d nuclei and clear or granular cytoplasm containing fine fat droplets.
The authors thank Drs. M. Kanbara and T. Kojima of the Department of Orthopedics, Ohita Red Cross Hospital, for helpful clinical comments. The authors also thank Tim Cornish for English language revision, and T. Umezu for technical assistance.
M y x o i d and R o u n d Cell L i p o s a r c o m a s
REFERENCES 1. DuBoulay, CEH (1985): Immunohistochemistry of soft tissue tumours. J Pathol 146:77-94. 2. Fletcher JA, Kozakewich HP, Hoffer FA, Lage JM, Weidner N, Tepper R, Pinkus GS, Morton CC, Corson JM (1991): Diagnostic relevance of clonal cytogenetic aberrations in malignant softtissue tumors. N Engl J Med 324:436-442. 3. Sandberg AA, Turc-Carel C, Gemmill RM (1988): Chromosomes in solid tumors and beyond. Cancer Res 48:1049-1059. 4. Turc-Carel C, Dal Cin P, Limon J, Rao U, Li FP, Corson JM, Zimmerman R, Parry DM, Cowan JM, Sandberg AA (1987): Involvement of chromosome X in primary cytogenetic change in human neoplasia: Nonrandom translocation in synovial sarcoma. Proc Natl Acad Sci USA 84:1981-1985. 5. Turc-Carel C, Aurias A, Mugneret F, Lizard S, Sidaner I, Volk C, Thiery JP, Olschwang S, Philip I, Berger MP, Philip T, Lenoir GM, Mazabraud A (1988): Chromosomes in Ewing's sarcoma. I. An evaluation of 85 cases and remarkable consistency of t(11;22)(q24;q12). Cancer Genet Cytogenet 32:229-238. 6. Whang-Peng J, Triche TJ, Knusten T, Miser J, Douglas EC, Israel MA (1984): Chromosome translocation in peripheral neuroepithelioma. N Engl J Med 311:584-585. 7. Sheng W-W, Soukup S, Ballard E, Gotwals B, Lampkin B (1988): Chromosomal analysis of sixteen rhabdomyosarcomas. Cancer Res 48:983-987. 8. Turc-Carel C, Limon J, Dal Cin P, Rao U, Karakousis C, Sandberg AA (1986): Cytogenetic studies of adipose tissue tumors. II. Recurrent reciprocal translocation t(12;16)(q13;p11) in myxoid liposarcomas. Cancer Genet Cytogenet 23:291-299. 9. Ohjimi Y, Iwasaki H, Ishiguro M, Ohgami A, Yoshitake K, Fujita C, Kikuchi M, Shinohara N, Kaneko Y: Myxoid liposarcoma with t(12;16)(q13;p11). Possible usefulness of chromosome analysis in a poorly differentiated sarcoma. Pat Res Pract (in press). 10. Limon J, Dal Cin P, Sandberg AA (1986): Application of longterm collagenase disaggregation for the cytogenetic analysis of human solid tumors. Cancer Genet Cytogenet 23:305-313. 11. ISCN (1985): An International System for Human Cytogenetic Nomenclature, Harnden DG, Klinger HP (eds.); published in collaboration with Cytogenet Cell Genet (Karger, Basel, 1985); also in Birth Defects: Original Article Series, Vol. 21, No. 1 (March of Dimes Birth Defects Foundation, New York, 1985). 12. Streekantaiah C, Karakousis CP, Leong SPL, Sandberg AA (1991): Trisomy 8 as a nonrandom secondary change in myxold liposarcoma. Cancer Genet Cytogenet 51:195-205.
117
13. Becher R, Wake N, Gibas Z, Ochi H, Sandberg AA (1984): Chromosome change in soft tissue sarcomas. J Natl Cancer Inst 72:823-831. 14. Karakousis CP, Dal Cin P, Turc-Carel C, Limon J, Sandberg AA (1987): Chromosomal changes in soft-tissue sarcomas. A new diagnostic parameter. Arch Surg 122:1257-1260. 15. Smith S, Reeves BR, Wong L (1987): Translocation t(12;16) in a case of myxoid liposarcoma. Cancer Genet Cytogenet 26:185-186. 16. Bridge JA, Shaffer B, Neff JR, Sanger WG, Moran J (1988): A complex translocation involving chromosomes 12 and 16 in a metastatic myxoid liposarcoma. Cancer Genet Cytogenet 34:119-120. 17. Molenaar WM, DeJong B, Buist J, Idenburg VJS, Seruca R, Vos AM, Hoekstra HJ (1989): Chromosomal analysis and the classification of soft tissue sarcomas. Lab Invest 60:266274. 18. Eneroth M, Mandahl N, Helm S, et al. (1990): Localization of the chromosomal breakpoints of the t(12;16) in liposarcoma to subbands 12q13.3 and 16p11.2. Cancer Genet Cytogenet 48:101-108. 19. Fujii Y, Matui Y, Nakagawa Y, Hongo T, Igarashi Y, Yamada M, Nakagome Y, Tobayama S (1989): Translocation t(12;16)(q13;p11) in myxoid liposarcoma of a child and implication of the human int-1 gene in tumorigenesis. Jpn J Cancer Res 80:958-962. 20. Orndal C, Mandahl N, Rydholm A, Nilbert M, Helm S, Akerman M, Mitelman F (1990): Chromosomal evolution and tumor progression in a myxoid liposarcoma. Acta Orthop Scand 61:99-105. 21. Mertens F, Johansson B, Mandahl N, Helm S, Bennet K, Rydholm A, Will6n H, Mitelman F (1987): Clonal chromosome abnormalities in two liposarcomas. Cancer Genet Cytogenet 28:137-144. 22. Walter TA, Decker HJH, Leong SPL, Sandberg AA (1988): A case of myxoid liposarcoma with translocation t(12;16) as the only abnormality. Cancer Genet Cytogenet 34:117-118. 23. Mugneret F, Lizard S, Aurias A, Turc-Carel C. (1988): Chromosomes in Ewing's sarcoma II. Nonrandom additional changes, trisomy 8 and der(16)t(1;16). Cancer Genet Cytogenet 32:239-245. 24. Sandberg AA (1986): The chromosomes in human leukemia. Semin Hematol 23:201-217. 25. Enzinger FM, Weiss SW (1988): Soft Tissue Tumors, 2nd Ed., C.V. Mosby, St. Louis, p. 357.
ABSTRACT: We performed a cytogenetic study of short-term cultures from fresh surgical specimens obtained from four patients with liposarcoma. Myxoid liposarcomas (cases 1-3) were associated with a specific translocation between chromosomes 12 and 16. Trisomy 8, a nonrandom secondary aberration in myxoid liposarcoma, was observed in the third case as the only additional change. Round cell liposarcoma (case 4) showed complex chromosomal aberrations affecting chromosomes 1, 2, 5, 6, 7, 13, 14, 17, 19, and 22. Neither band 12q13 nor 16pl I was visibly rearranged. Three subgroups of liposarcomas are proposed. The first group is characterized by t(12;16)(q13;p11), the second group by ring chromosomes, telomeric associations, and giant markers, and the last by complex numerical and structural aberrations.
INTRODUCTION Differential diagnosis in soft tissue tumors is often very difficult for both clinicians and pathologists. Special stains, immunohistochemistry, and electron microscopy have been introduced as useful tools for pathologic studies, but poorly differentiated sarcomas remain the most difficult tumors to diagnose [1]. Cytogenetic study is now recognized as a useful technique for diagnosis and estimation of prognosis for blood malignancies. Recently, cytogenetic studies have also proved useful for analysis of solid tumors [2, 3]. Apparently diagnostic chromosome rearrangements have been described in several varieties of malignant soft tissue tumors. These rearrangements include t(X;18)(p11.2;q11.2) in synovial sarcoma [4], t(11;22)(q24;q11.2-12) in Ewing's sarcoma [5] and peripheral primitive neuroectodermal tumors [6], t(2;13)(q35-37;q14) in alveolar rhabdomyosarcoma [7], and t(12;16)(q13;p11) in myxoid liposarcoma [8]. We report the results of chromosome analysis in three cases of myxoid liposarcoma and one case of round cell liposarcoma.
MATERIALS AND METHODS Clinical Data All patients were Japanese, and clinical data of patients are summarized in the Table 1. Case 1 was detailed in another article [9]. From the Department of Pathology (Y. 0., H. I., M. I., A. 0., C. F., K. Y., M. K.I, Fukuoka University School of Medicine, Fukuoka, Laboratory Medicine (Y. K.), Saitama CancerCenter, Saitama, and Department of Orthopedics (N. S.), National Fukuoka Central Hospital, Fukuoka, Japan. Address reprint requests to: Yuko Ohjimi, M.D., Department of Pathology, Fakuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka 814-01, Japan. Received February 21, 1992; accepted May 14, 1992.
Surgical Specimens Tumor tissues were divided immediately into four parts and processed for light and electron microscopy, frozen sectioning, and short-term culturing. Light Microscopy The tumor tissues were fixed in 10% formalin or B-5 fixative. Paraffin sections prepared from these samples were stained with H&E and alcian blue with or without prior hyaluronidase digestion. To detect fat in the tumor cells, cryostat sections were prepared from the frozen tumor tissue and stained with oil red-O. Electron Microscopy The fresh surgical specimens were fixed in 3.0% glutaraldehyde in 0.1 M cacodylate buffer, postfixed in 1.0% osmium tetroxide, dehydrated in ethyl alcohol, and embedded in Epok 812 (Nagase & Co. Osaka, Japan). The sections were cut with a Reichert ultramicrotome, stained with uranylacetate and lead citrate, and examined with a JEM 100 CX-II electron microscope (Jeol, Tokyo, Japan). Cytogenetic Investigation Minced fragments of the tumor tissue were suspended in 200 U/ml type II collagenase (Worthington Biochemical Corporation, Freehold, NJ) for 16 hours [10]. After digestion, the tumor cells were washed and resuspended in the culture medium in glass flasks (TD40, Ikemoto Scientific Technology, Tokyo, Japan), and then incubated at 37°C in a humidified atmosphere containing 5% CO 2 and 5% 02. The culture medium consisted of ES medium "Nissui" (Nissui Pharmaceutical, Tokyo, Japan), supplemented with 10-17% fetal calf serum (HyClone Laboratories, Logan, UT) and 0.3% glutamine (Nissui Pharmaceutical Co., Tokyo, Japan). Colcemid (Life Technologies, Gaithersburg, MD) 111
© 1992 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York, NY 10010
Cancer Genet Cytogenet 64:111-117 (1992) 0165-4608/92/$05.00
112
Table 1
Y. Ohjimi et al.
Clinical and cytogenetic data of liposarcoma patients
Case no.
Primary Age/ tumor sex site
Histologic type
RT or Source CT
Tumor size (cm)
Prognosis (mo)
10 × 10
No. of
Culture metaphases (days) analyzed
1.SFT8925 57/F
R thigh
Myxoid LS
P
No
18, AWD
3
2.SFT9019 39/F 3.SFT9034 35/M
L thigh L thigh
Myxoid LS Myxoid LS
P R
No 5 x 6 11, AWD No 2 × 1.5 × 1 6, AWD
2 5
4.SFT9002 40/M
R thigh
Round cell LS
P
No
2
10 x 10
17, AWD
5 2 33 10 1 8 12
Karyotype 46,XX,t(12;16)(q13;p11) 46,XX 46,XX,t(12;16)(q13;p11) 47,XY, + 8,t(12;16)(q13;p11) 46,XY,t(12;16)(q13;p11) 46,XY 56-62,XY, + 1, + 2, + 5, + 6, + 7 , - 13,- 13,- 14, der(17)t(17;?)(q25;?), + der(17),+ 19,- 22, +9-15 mar
Abbreviations: LS, liposarcoma;P, primary tumor; R, recurrent tumor; RT, radiationtherapy; CT, chemotherapy; AWD.alive without disease.
was added in the last 2 hours at a final concentration of 0.02 /xg/ml. Cells were harvested by trypsinization. After hypotonic shock in 0.075 M KC1 solution for 2 0 - 3 0 minutes, cells were fixed gradually by addition of Carnoy's solution. Cells were spread on moisture slide glasses and dried at room temperature for 2 days. Trypsin-Giemsa b a n d i n g stain was performed. The karyotypes were described according to the short system of the International System for H u m a n Cytogenetic Nomenclature [11].
RESULTS Light Microscopy In case 1, H&E stain showed proliferation of round and polygonal cells with atypical hyperchromatic nuclei. Hypercellular areas with a few mitotic figures were observed. The tumor cells appeared immature, showing no obvious lipoblastic differentiation. Fine networks of capillary blood vessels forming an alveolar pattern were evident in the stroma (Fig. 1A and B). Alcian blue stain demonstrated an a b u n d a n t mucous substance in the intercellular matrix; the substance was digested by hyaluronidase. Oil red-O stain showed only small fat droplets in the cytoplasm of the original tumor. Cases 2 and 3 showed typical histologic features of myxold liposarcoma, with proliferating lipoblasts, delicate plexiform capillary patterns, and an a b u n d a n t myxoid matrix (Fig. 1C and D). Oil red-O stain showed a b u n d a n t and large fat droplets in the cytoplasm. In case 4, round tumor cells with pale or slightly eosinophilic and somewhat vacuolated cytoplasm were arranged in sheetlike pattern with a fine vascular network. Many mitotic figures were apparent among the tumor cells. Oil red-O stain on frozen sections showed small fat droplets in
tumor cells. The histologic feature was indicative of poorly differentiated found cell liposarcoma (Fig. 2).
Electron Microscopy In case 1, the tumor cells generally had few and poorly developed organelles, although some cells contained dilated rough endoplasmic reticulum. The cells possessed a few small n o n m e m b r a n e - b o u n d lipid droplets (Fig. 1E). In cases 2 and 3, the electron microscopic findings were almost the same. Many tumor cells had one or more nonm e m b r a n e - b o u n d lipid droplets of variable size and density and little or no endoplasmic reticulum. The nuclei were large and deeply indented and located at the periphery of the cell (Fig. 1F). In case 4, some tumor cells had lipid droplets, large nuclei with p r o m i n e n t nucleoli, and sparse organelles. There were no desmosomes or other cellular attachments.
Cytogenetic Analysis Results of cytogenetic analysis are summarized in Table 1. Case 1. Of the seven G-banded metaphases analyzed, five
showed the karyotype 46,XX,t(12;16)(q13;p11), with the translocation as the only chromosome change (Fig. 3, left). One cell showed an apparently normal karyotype, 46,XX, and the other showed a karyotype of 4 5 , X X , - 1 5 , which was considered a random missing chromosome. Case 2. Thirty-three analyzable metaphases were obtained.
All showed the clonal (q13;p11) (Fig. 3, center).
karyotype
46,XX,t(12;16)
Case 3. Nineteen metaphases were adequate for cytoge-
netic analysis. Ten of them showed 47,XY,+8,t(12;16)
Figure 1 (A) Delicate capillary network and abundant my×old matrix (case 1, H&E, original magnification x 130). (B) Proliferation of round to polygonal tumor cells with hyperchromatic nuclei (case 2, H&E, original magnification x 515). (C) Delicate plexiform capillary patterns and abundant myxoid matrix are evident (case 3, H&E, original magnification x 130). (D) Proliferating lipoblasts and delicate capillaries (case 3, H&E, original magnification x 515). (E) Electron micrograph of case 1 showing dilated rough endoplasmic reticulum and a few fat droplets (original magnification × 8,550). (F) Electron micrograph of case 3 showing tumor cell with nonmembrane-bound lipid droplets and little endoplasmic reticulum (original magnification x 9,150).
114
A
B Figure 2
Histologic pictures of round cell liposarcoma (case 4). (A) Fine vascular networks and proliferation of round cells (H&E, original magnification × 130). (B) Round tumor cells with pale or slightly eosinophilic cytoplasm (H&E, original magnification x 515).
(q13;p11), one s h o w e d 46,XY,t(12;16)(q13p11), and the other eight showed an a p p a r e n t l y normal karyotype, 46,XY (Fig. 3, right). Case 4. Twelve metaphases were karyotyped. There were many numerical and structural aberrations. Although no two metaphases showed exactly the same karyotypes, many c o m m o n aberrations were noted. A representative karyotype was 5 6 - 6 2 , X Y , + 1 , + 2 , + 5 , + 6 , + 7 , - 1 3 , - 1 3 , - 1 4 , der(17)t(17;?)(q25;?), + tier(17), + 1 9 , - 22, + 9 - 1 5 m a r (Fig. 4).
DISCUSSION Soft tissue (mesenchymal) n e o p l a s m s are rare, accounting for less than 1% of h u m a n malignancy. Unless histopathologists work in large cancer centers, they see few of these tumors. Moreover, undifferentiated or poorly differentiated soft tissue tumors often lack any specific cellular feature.
Y. Ohjimi et al. Recently, cytogenetic analysis of soft tissue tumors has become a valuable tool in diagnosing and defining these tumors and even in providing clues to their histogenesis [2, 3]. Some of these tumors are now k n o w n to have specific c h r o m o s o m e abnormalities. In 1986, Turc-Carel et al. [8] first described a t(12;16)(q13;p11) as a characteristic aberration of m y x o i d liposarcoma, w h i c h is n o w considered one of the tumorspecific c h r o m o s o m e aberrations. In our case 1, pathologic findings such as fine capillary networks, r o u n d to polygonal tumor cells with atypical h y p e r c h r o m a t i c nuclei, and myxold stroma were suggestive of m y x o i d liposarcoma. Ultrastructural examination and oil red-O fat stain failed to demonstrate any evidence of lipoblastic differentiation in the tumor cells, however. Chromosome analysis of the tumor cells in case 1 showed a reciprocal t(12;16)(q13;p11). The existence of t(12;16) confirmed the diagnosis of m y x o i d liposarcoma in this case. Cases 2 and 3 showed typical histologic features of myxoid l i p o s a r c o m a and t(12;16)(q13;p11). Case 3 exhibited an additional numerical aberration: trisomy 8. In the available literature, 28 cases of cytogenetically analyzed m y x o i d and related liposarcomas have been reported [2, 8, 12-22]. We report three additional cases of m y x o i d liposarcoma, including two primary tumors and one recurrent tumor, i.e., 32 tumors in 31 patients (Table 2), including 14 primary, 12 recurrent, and 5 metastatic tumors. The t(12;16)(q13;p11) was apparent in 20 (61%) of the tumors. The same translocation was evident in 12 (85%) of the primary tumors. Aberrations with breakpoints in 12q13 and 1 6 p l l were noted in 26 (81%) and 24 (75%) of the tumors. Although some of the m y x o i d liposarcomas did not show both breakpoints, molecularilevel changes cannot be ruled out for both bands [18]. Eneroth et al. [18] and Streekantaiah et al. [12] reported that trisomy 8 existed as a n o n r a n d o m secondary change in m y x o i d liposarcoma. Mugneret et al. [23] reported that trisomy 8 was observed in 44% of Ewing's sarcomas as well. Trisomy 8 is a c o m m o n secondary numerical abnormality in both chronic and acute m y e l o i d leukemias and indicates a poor prognosis [24]. Case 4 was a r o u n d cell liposarcoma with complex numerical and structural aberrations. Numerical aberrations were evident in chromosomes 1, 2, 5, 6, 7, 13, 14, 17, 19, and 22, with 9 - 1 5 unidentified marker chromosomes and one or two markers derived from chromosome 17. That no two metaphases showed exactly the same karyotype suggests cytogenetical instability in this tumor. Two other cases of cytogenetically analyzed round cell liposarcoma exist [17]; both exhibit c o m p l e x chromosomal aberrations. All three r o u n d cell liposarcomas affected chromosomes 5 and 7 and two affected chromosomes 2, 6, 10, 14, 17, but none of them showed t(12;16)(q13;p11). Therefore, round cell liposarcomas a p p a r e n t l y are cytogenetically different from m y x o i d liposarcomas. Round cell liposarcomas are closely related to myxoid liposarcoma, and most have histologic transitions toward the m y x o i d type. In some instances, the initial tumor is of the m y x o i d type and the recurrent or metastatic tumor is of the round cell type [25]. Four cases of " m i x e d - t y p e "
Myxoid and Round Cell Liposarcomas
115
/
/
1
/
/
/
Figure 3 Partial karyotypes of case 1 (left), case 2 (center), and case 3 (right) showing t(12;16)(q13;p11).
liposarcoma, which probably means the transitional form of liposarcoma, have been reported [2, 18, 21]. Three of them showed t(12;16), and the other showed a complex karyotype without the specific t(12;16). The investigators questioned any cytogenetical relation existed between myxoid, mixed, and round cell liposarcomas. There are three possible answers to this question; First, although histologic transition is observed between myxoid and round cell liposarcomas, there are no cytogenetic transitions. Therefore, myxoid and "mixed" liposarcomas belong to the same group characterized by t(12;16), and round cell liposarcomas belong to another cytogenetic group charac-
terized by complex karyotypes. The second possibility is that round cell liposarcomas have truly mixed abnormalities, including t(12;16)(q13;p11), and that t(12;16)(q13;p11) is masked by complex aberrations. Third, myxoid areas of mixed type have t(12;16)(q13;p11), and round cell areas of mixed type have complex aberrations. We agree with the last answer, because three different clones were found in different parts of one myxoid liposarcoma [20]. We propose the existence of at least three cytogenetic groups of liposarcoma. The first group is characterized by a special translocation, i.e., t(12;p16)(q13;p11). This group has the histologic features of myxoid liposarcoma, and the
Figure 4. Karyotype of case 4 showing complex aberrations: 60,XY,+1,+2,+5,+6,+7,+8,-13,-13,-14, - 17, - der(17)t(17;?)(q25;?) × 2, + der(17;?)(q;?),+ 19, - 22, - 22, + 12mar (arrows point to abnormal chromosomes).
"11 I
!1i 8
7
6
4
5
10
11
12
16
i7
18
3
2
il 9
f
If 14
13
15
f 19 ¸
fl
If 2O
21
22
X
Y
116
Y. Ohjimi et al.
Table 2
Myxoid and related lipasarcomas
Histologic Ref type
Source
RT or CT R
[12] [12]
My My
R P
{121 [12]
My My
R R
[13] [81 [8] [8] [8]
My My My My My
M P P P R
[14]
My
R
[141 [15] [16] [17]
My My My My
R M M R
[171
My My
a)M b)M
[18]
My
P
[18]
My
P
[19] [20]
My My
R PA P B1 P B2
118] [181
Mix Mix Mix Mix
[2] [21]
P B3 P P P
Primary tumor site
Karyotype
R thigh R groin
46,XX,t(12;16)(q13;p11} 46,XY,t(12;16)(q13;p11) 47,XY, + 8,t(12;16)(q13;p11/ R R pelvis 48,XX, + 8,t(11;17)(q13;q21),t(12;16](q13;p11},+ 19 R and C L thigh 46,XY 46,XY,t(12;16)(q13;p11) 46,XY 48,XY,- 9, + 10, - 11, + der(11)t(11;?)(p15;?),t(12;16)(q13;p11),- 17, + 1-3mar R or C R thigh der(12}, 10% of analyzed cells R thigh 46,XX,t(3;15)(p23;q15),t(12;16)(q13;p11) R leg 46,XY,del(6)(q21),t(12;16)(q13;p111 Retroperitoneum 46,XX,t(12;16)[q13;p11) C R thigh 46,XY, + 1,der(1)t(1;12;16)(p11;q13;p11),- 16, der(12)t(1;12;16)(p11;q13;p11) Retroperitoneum del(1)(q11),del(2)(p11),del(3)(p21),t(12;19)(q13;p or q13)[loss der(12)], 6q+,11p+,12p+ Leg 46,XY/46,XY,t(12;16)(q13;p11) ? + der(1)t(1;?)(p11 or p13;?), + 8,t(12;16)(q13;p11),- 16 ? t(12;16;16}(pllq21;p11;q12) Thigh 47,XY,t(1;12)(q42;q13),t(1;16)(q22;p11), + 8 47,XY,+ 8 46,XY,t(11;12}(p15;q13) Thigh 47,XY,- 2,del(6}(q11),t(11;12)(p15;q13),der(15)t(15;?)(q24;?), der(21)t(1;21)(q12;q22), + marl(del(2)?), + mar2 No Thigh 46,XY,der(12)t(12;16)(q13;p11),der(16)t{12;16)(q22;p11) 46,XY No Thigh 46,XX,t(12;16](q13;p11) 46,XX,i(7q),t(12;16)(q13;p11) R and C Retroperitoneum 46,XX,t(12;16)(q13;p11), near tetraploid R thigh 46,XY,t(12;16)(q13;p11) 46,XY,t(12;16)(q13;p11) 46,XY,t(12;16)(q13;p11) 46,XY, + der(1)t(1;16)(p11;p11),t(12;16)(q13;p11),- 16 46,XY 46,XY, + der(1)t(1;12)(p11;q13),der(12)t(12;16)(q13;p11),- 16 47,XY, + 8,t(12;16)(q13.3;p11.2) No Thigh No Thigh 46,XY,t(12;16)(q13.3;p11.2) Thigh Absence of t(12;16); complex karyotype R leg 46,XY,t{12;16)(q13;p11) ?
[121
My+PD
R
]22] My+WD
P
]14] WD+My [14] WD+ My ]14] WD+My
P R R
L thigh
47,XY, + 8,t(12;16)(q13;p11) 94,XXYY, + 8, + 8,t(12;16)(q13;p11),t(12;16)(q13;p11) endoredup|ication with t(12;16) Rthigh 46,XY,t(12;16)(q13;p11} 46,XY Retroperitoneum 11q + ,ring,tas,giant markers Retroperitoneum Near triploid, UMs Retroperitoneum Ring,tas,giant markers
Abbreviations: My, myxoid;Mix, mixed; PD, poorly differentiated;WD, well differentiated;tas, telomeric associations;UMs, unidentifiedmarkers; other abbreviationsas in Table 1. tumor frequently occurs in the thigh. In 1986, Turc-Carel et al. [8] suggested that tumors with this translocation might have a better prognosis. The second group consists of liposarcomas with ring chromosomes, telomeric associations, and giant markers [14]. They have a tendency to occur in the retroperitoneum and histopathologically exhibit the features of n o n m y x o i d (differentiated or pleomorphic) lipo* sarcomas. The third group is characterized by complex numerical and structural aberrations. The tumors often
have the histologic feature of r o u n d cell liposarcoma, characterized by r o u n d nuclei and clear or granular cytoplasm containing fine fat droplets.
The authors thank Drs. M. Kanbara and T. Kojima of the Department of Orthopedics, Ohita Red Cross Hospital, for helpful clinical comments. The authors also thank Tim Cornish for English language revision, and T. Umezu for technical assistance.
M y x o i d and R o u n d Cell L i p o s a r c o m a s
REFERENCES 1. DuBoulay, CEH (1985): Immunohistochemistry of soft tissue tumours. J Pathol 146:77-94. 2. Fletcher JA, Kozakewich HP, Hoffer FA, Lage JM, Weidner N, Tepper R, Pinkus GS, Morton CC, Corson JM (1991): Diagnostic relevance of clonal cytogenetic aberrations in malignant softtissue tumors. N Engl J Med 324:436-442. 3. Sandberg AA, Turc-Carel C, Gemmill RM (1988): Chromosomes in solid tumors and beyond. Cancer Res 48:1049-1059. 4. Turc-Carel C, Dal Cin P, Limon J, Rao U, Li FP, Corson JM, Zimmerman R, Parry DM, Cowan JM, Sandberg AA (1987): Involvement of chromosome X in primary cytogenetic change in human neoplasia: Nonrandom translocation in synovial sarcoma. Proc Natl Acad Sci USA 84:1981-1985. 5. Turc-Carel C, Aurias A, Mugneret F, Lizard S, Sidaner I, Volk C, Thiery JP, Olschwang S, Philip I, Berger MP, Philip T, Lenoir GM, Mazabraud A (1988): Chromosomes in Ewing's sarcoma. I. An evaluation of 85 cases and remarkable consistency of t(11;22)(q24;q12). Cancer Genet Cytogenet 32:229-238. 6. Whang-Peng J, Triche TJ, Knusten T, Miser J, Douglas EC, Israel MA (1984): Chromosome translocation in peripheral neuroepithelioma. N Engl J Med 311:584-585. 7. Sheng W-W, Soukup S, Ballard E, Gotwals B, Lampkin B (1988): Chromosomal analysis of sixteen rhabdomyosarcomas. Cancer Res 48:983-987. 8. Turc-Carel C, Limon J, Dal Cin P, Rao U, Karakousis C, Sandberg AA (1986): Cytogenetic studies of adipose tissue tumors. II. Recurrent reciprocal translocation t(12;16)(q13;p11) in myxoid liposarcomas. Cancer Genet Cytogenet 23:291-299. 9. Ohjimi Y, Iwasaki H, Ishiguro M, Ohgami A, Yoshitake K, Fujita C, Kikuchi M, Shinohara N, Kaneko Y: Myxoid liposarcoma with t(12;16)(q13;p11). Possible usefulness of chromosome analysis in a poorly differentiated sarcoma. Pat Res Pract (in press). 10. Limon J, Dal Cin P, Sandberg AA (1986): Application of longterm collagenase disaggregation for the cytogenetic analysis of human solid tumors. Cancer Genet Cytogenet 23:305-313. 11. ISCN (1985): An International System for Human Cytogenetic Nomenclature, Harnden DG, Klinger HP (eds.); published in collaboration with Cytogenet Cell Genet (Karger, Basel, 1985); also in Birth Defects: Original Article Series, Vol. 21, No. 1 (March of Dimes Birth Defects Foundation, New York, 1985). 12. Streekantaiah C, Karakousis CP, Leong SPL, Sandberg AA (1991): Trisomy 8 as a nonrandom secondary change in myxold liposarcoma. Cancer Genet Cytogenet 51:195-205.
117
13. Becher R, Wake N, Gibas Z, Ochi H, Sandberg AA (1984): Chromosome change in soft tissue sarcomas. J Natl Cancer Inst 72:823-831. 14. Karakousis CP, Dal Cin P, Turc-Carel C, Limon J, Sandberg AA (1987): Chromosomal changes in soft-tissue sarcomas. A new diagnostic parameter. Arch Surg 122:1257-1260. 15. Smith S, Reeves BR, Wong L (1987): Translocation t(12;16) in a case of myxoid liposarcoma. Cancer Genet Cytogenet 26:185-186. 16. Bridge JA, Shaffer B, Neff JR, Sanger WG, Moran J (1988): A complex translocation involving chromosomes 12 and 16 in a metastatic myxoid liposarcoma. Cancer Genet Cytogenet 34:119-120. 17. Molenaar WM, DeJong B, Buist J, Idenburg VJS, Seruca R, Vos AM, Hoekstra HJ (1989): Chromosomal analysis and the classification of soft tissue sarcomas. Lab Invest 60:266274. 18. Eneroth M, Mandahl N, Helm S, et al. (1990): Localization of the chromosomal breakpoints of the t(12;16) in liposarcoma to subbands 12q13.3 and 16p11.2. Cancer Genet Cytogenet 48:101-108. 19. Fujii Y, Matui Y, Nakagawa Y, Hongo T, Igarashi Y, Yamada M, Nakagome Y, Tobayama S (1989): Translocation t(12;16)(q13;p11) in myxoid liposarcoma of a child and implication of the human int-1 gene in tumorigenesis. Jpn J Cancer Res 80:958-962. 20. Orndal C, Mandahl N, Rydholm A, Nilbert M, Helm S, Akerman M, Mitelman F (1990): Chromosomal evolution and tumor progression in a myxoid liposarcoma. Acta Orthop Scand 61:99-105. 21. Mertens F, Johansson B, Mandahl N, Helm S, Bennet K, Rydholm A, Will6n H, Mitelman F (1987): Clonal chromosome abnormalities in two liposarcomas. Cancer Genet Cytogenet 28:137-144. 22. Walter TA, Decker HJH, Leong SPL, Sandberg AA (1988): A case of myxoid liposarcoma with translocation t(12;16) as the only abnormality. Cancer Genet Cytogenet 34:117-118. 23. Mugneret F, Lizard S, Aurias A, Turc-Carel C. (1988): Chromosomes in Ewing's sarcoma II. Nonrandom additional changes, trisomy 8 and der(16)t(1;16). Cancer Genet Cytogenet 32:239-245. 24. Sandberg AA (1986): The chromosomes in human leukemia. Semin Hematol 23:201-217. 25. Enzinger FM, Weiss SW (1988): Soft Tissue Tumors, 2nd Ed., C.V. Mosby, St. Louis, p. 357.
