Subgroups of uterine leiomyomas based on cytogenetic analysis

Subgroups of Uterine Leiomyomas Based on cytogenetic Analysis JIE H\J, MD, Chromosomes

AND

URVASHI

from 39 cases of benign

studied. Consistent

chromosomal

15 cases (38.5%). Abnormalities with or without additional

SURTI,

uterine leiomyomas

abnormalities

14 were obsewed mosome

involving

and nonreciprocal

in

present.

X, 5, and 14 as well as X, 3. and

1, deletion

translocation

involving

chromosome

between chromosomes

were observed in one case each. Monosomy

3,

14 and 15

22, with a derived chro-

mosome 14. was observed in one case. Trisomy 7 was also identified in one case. The structural

and numeric

abnormalities

involved

chromosome+; X. 1, 2, 3, 4, 5, 6, 7, 9, 10, 12, 13, 14, 15, and 22. A normal 46,Xx stem line with one or two abnormal cells was observed in 20 cases. Olnly normal karyotypes were obtained in the remaining four cases. A review of the literature study indicate that uterine leiomyomas groups

based

on cytogenetic

and the results of our

may be divided

analysis.

Cytogenetic

7 was detected in five

in one case each. Insertion of a portion of chro-

4 to chromosome

hours. The cells were cultured in (:hang mrdium at -1 to 10 days after growth was cstablishcd

HUM

PATHOL

Analysis

For this study, 32 tumors were prepared fi~r tissue culture. Thirty-nine of 42 tumors had successful growdl. Normal tissue adjacent to the tunlor area in 26 cases was also prepared fol culture as a c’onlrol. At least 20 mrtaphase cells \vtbre evamiried for each case. Q-handing was the primary method used for determining rhr chromosomal abnormalities. (;- and C-banding were used as complementary methods. Cytogenetic findings were recorded according 10 the International System for HuMuscle-spetcifiit- anridrsmin man Cyrogenetic Nomenclature.” antibody &IS used to identify the nature of culturecl cells in seven cases.

into eight 22:1009-

RESULTS

1016. Copyri:e;ht ~1 1991 by W.B. Saunders Company

Nonrandom chromosomal changes have been identified in a number of benign tumors, such as lipoInas,‘.’ meningiomas,” and mixed salivary gland tumors.’ Recently, several groups have reported that clonal chromosomal rearrangements also exist in benign uterIt has been suggested that reciprocal ine leiomyomas.‘~” t ranslocation between chromosomes 12 and 14 and interstitial deletion of chromosome 7 may characterize two subgroups of uterine leiomyomas.‘S”’ A question of obvious interest is whether there are any other subgroups with characteristic chromosomal changes. We report the cytogenetic results of 39 uterine leiomyomas from 39 patienrs.

MATERIALS

;md hanestcsd

12 and 14

changes were found

cases; in three cases (7.6%), this was the only abnormality tramlocations

were

were detected in

involving chromosomes

chromosomal

five cases (12.8%). Deletion of chromosome Complex

PHD

AND METHODS

Tissue Culture Saniple~ of uterine leiomyomas were obtained from patiellrs who underwent operations at Magee-M’omens Hospital. Thq were t\.pical leionlyomas proven by histopathologic ex.miination. i‘he 11-41 tunlor tissue was finely minced, then en/ymatically disaggregated with 0.125% trypsin (Worthington Kiochrmical ~~~or-p. Freehold, NJ) for 30 minutes and with 0.6% c~ollagenase [Sigma (:hemical (:o, St Louis, MO) for 4 to ti

1009

Clonal chromosomal rearrangements were observed in 38.5% of cases (15 of 39). normal 46,Xx stem lines with one or two different abnormal cells were observed in 51.3% of cases (20 of 39). and normal karyotypes were observed in the remaining 1~0.2Y0 of cases (four of 39). The structural abnormalities involved chromosomes X, 1, 2, 3, 4, 5, 6, 7. 8, 9, 10, 11, 12, 13, 14, 15, and 22. Most frequently, they involved chromosomes 1, 3, 7, 12, and 14. All of the seven cases stained for desmin showed positive staining. translocations between Ca.w uo. I. Reciprocal chromosomes 12 and 14, as well as between chromosomes 6 and 10, were found in 25 of 26 metaphase cells studied. The karyotype is 46,XX,t(t’S; lO)(pl l;q23), t(l2; 14)(q15;q23) (Fig 1). Casv no. 2. This tumor showed a karyotype 15,Xx,14,-22,der( 14) in all 30 cells examined. The origin of the abnormal chromosome 14 could not be determined (Fig 1). Casu ~ZO.3. In this case, a complex chromosomal rearrangement was seen in 24 of 25 cells analyzed, with a modal karyotype of 46,XX,t[inv(12)1(~13q 15); 141 [p13;q24] (Fig 1). Cusp no. 4. A complex translocation involving chromosomes 12, 13, and 14 accompanied by interstitial deletion of chromosome 7 and inversion in chromosome 1 was observed in all 30 cells analyzed. Karyotype 4~,XX,i~~v(l)(p22~1251,del(7~(ql lq22).t(14; 13: 1-l) (ql5;q3 1 ; q22) was found in 28 cells (Fig 2 I. In addition to the above rearrangements. six cells sllowed monosomy 22. and two cells had trisomv 1, 8, and 10. Four cells were hypodiploid. (One cell was missing both chromosomes 2 1 and one chromosome 22. One c.ell was missing one chromosome 7 and one chromosome 2 1. Two cells could not be karyotvped.)

HUMAN PATHOLOGY

Volume 22, No. IO (October

1991)

FIGURE 1. Q-banded partial karyotypes of cases no. 1, 2, 3, 5, and 7. (Maanitication x 1.900.) Case no. 1: ml -= der(b)t(6:lOj(pl l:q24): m2 = der(10)t(6;10)(pll;q24); m3 = der(12)t(12;14)(q15;q24); m4 = der(l4)t(l2;14)(ql5;q24). Case no. 2: m = der(14)t(14:?)(?;?). Case no. 3: ml = der(l2)(14c$er + 14a24::12a15 + 12o13::12a15 + 12aterI m2’= der(l4)(14ptkr -+ 14q24:: 12~1’3 1 12pter). Case no. 5: ml = der(3)(3pter -+ 3q27::14q22 + 14qter); m2 = der(14) (14pter + 14q22::Xq26 -+ Xqter); m3 = der(X)(Xpter -. Xq26::3q27 + 3qter). Caseno. 7:ml =der(l)(l3qter-+ 13q14 ::lp36 + Iqter); m2 = der(13)(q14); m3 = der(12)t(12;14)(q15;q24 m4 = der(14)t(12;14)(q15;q24 m5 = der(2)ins(2)?: m6 = ?

1010

CYTOGENETICS

OF UTERINE LEIOMYOMAS

(Hu & Surti)

GHROMOSOME BREAKPOINTS

REARRANGEMENTS OBSERVED

-

ml

1

ml 7qll 7q11-

7q22

7q22---FIGURE 2. Marker chromosomes in case no. 4: G-. C-, and Q-banding. (Magnification x 1,900.) m1 = inv(l)(p22q25); m2 = del(7) (q 1 lq22); m3 = der(12)t(12;13: 14)(q15:q31;q22): m4 = der (13)t (12; 13; 14)(q15;q31:q22); m5 = der (14)t(l2;13:14)(ql5.q3l;q22).

7

m2

7

m2

12 12q15IQ 12

12q15

<13

13q31

m3

0 13

m4 14

ti

14922-bB

8 1‘4

m5

1011

14

12

m5

<

1*= 12q15

HUMAN PATHOLOGY

Volume 22, No. 10 (October

C,ksc no. 5. Translocation involving chromosomes X, 3, and 14 was present in all 27 cells studied. The karyotype is 46,XX,t(X;3; 14)(q26;q27;q22) (Fig 1). Cuse ML 6. Translocations between chromosomes 2 and 4, as well as between chromosomes 12 and 14, were observed in all 32 cells studied. In addition, deletions involving chromosomes 3 and 7 were seen in all of the cells (Fig 3). Sixteen tumor cells showed karyotype 46,XX,t (2 ; 4) (~13 ; ~16) .del(3) (~13~14) .del(7) (q22). t(l2; ll)(q13;q22). Th e remaining 16 cells showed random chromosome loss. Figure 4 shows the ideogram of the rearrangements observed in this case. 46,Xx, -1,-2,-5,-l 3,+der Cuse 110. 7. Karyotype (1)(13qter+13q14::1p36+1qter),+der(2)ins(2;?) marker,t(l2; 13) (q15;q24) (Fig (q12;?).+de1(13)(ql4),+ 1) was found in 19 of 20 cells examined. The remaining one cell had 46,XX,del(l)(p36),t(12; 14)(q15;q24) karyotype (Fig 1). CUP 110. 8. 45,XX,-14,-15,t(l4;15)(q12?;p12?) was found in six of 20 cells. The remaining 14 cells were normal. Cusr IIO. 9. 46,XX,t(X; 5; 14)(q24?; p15; q24) was found in four of 24 cells. Case rm. 10. 46,XX,ins( 1; 4)(q32; q3 1 q35) was observed in four of 16 cells. Cuse 110. 11. 46,XX,de1(3)(p13p14) was found in four of 24 cells. CUSPWJ. 12. An interstitial deletion in chromosome region 7q was detected in four of 20 cells. The exact breakpoints could not be determined.

Koryotype of case no. 6 (Q-banding): nification k 1,900.)

FIGURE 3.

1991)

Cnse IW. 13. 46,XX,de1(7)(q22) was found in four of 20 cells. Cnse PLO.11. 46,XX,del(7)(q32) was found in five of 23 cells. Cuse ML 15. 47,Xx,+7 was observed in three of 20 cells. Twenty cases showed predominantly normal karyotypes, with one or two cells with nonclonal chromosomal abnormalities. These were thought to be random except for the chromosome break at the 12q15 region in a single cell in four different cases.

DISCUSSION Rearrangements involving 12ql3- 15 and 1 tq22-24 were found in five cases, including three cases with reciprocal translocations involving chromosomes 12 and 14. Deletions del(7)(ql lq22), de1(7)(q22), or de1(7)(q32) were observed in five cases. Among these cases, deletion 7 as the only change was identified in three cases. These two nonrandom changes have been suggested to represent two different subgroups of uterine leiomyomas. However, other subgroups may exist. (=hromosomal abnormalities involving 14q22-24 and other chromosomes, in which both chromosomes 12 looked normal, were observed in three cases in our study; this was also reported in two cases in previous studies.‘.’ Rearrangements involving chromosome 12 without affecting chromosome 14 were also found in five cases in previous

~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ (Mag-

CYTOGENETICS

OF UTERINE LEIOMYOMAS

(Hu & Surti)

s,udies,“,“.

II’, I’( Still other recurrent clonal c~l~~~or~~osor~~~~~ changes. such as translocation between chromowmes 1 and 2, haw been reported in five unrelated cases.” The fi~rmation of uterine leiomyomas may follow different cvtogenetic pathways. According to present and pre&us studies,‘.” chromosomal abnormaliltics of’ benign uterine leiomvomas may be divided into eight groups (TatA? 1): (;roup 1: Structural abnormalities involvirlg chromosomes 12 and 14. (;roup L’: Kearrangemerit invohirig c hroniosonie 12 and other chromosomes when c.hroniosonie 14 is normal. 11 and (;roup :<: (;hanges involving chron~osome other chromosomes. hut no1 involving chromosome I 2. Abnormalities involving c~tironiosoni~ ‘7 <;roLl[’ 4: (including interstitial and tcwninal deletion). lvith or without othw rearrmge-

4~16

4 --<

2~13

2

ml

2 2~13

--_(

4~16

4

m2

nlerlts.

involving chrom~~sonie\; 1 anti L) _. Qrrwtulal abi~ornial(;roup (i: Other nonrandom ities. c~hro~riosor~l~~l ;ibnornialities (;roiij~ i: Numeric (including 1risomies) Normal karyotypes. (;r-oLlp 8: ,411of the ~hronww)me breakpoints obwrvecl in our S!J cxses are shown in Fig 5. The breakage invol\w~ 17 chroniosonies and 4.5 regions. Sis&-four percxwt of breakpoints seem to coincide with [tic. lo(:ation~ of OII~~~gcne- ant1 i~~l~nuno~lohulin-asso~i~ite[l genes. Based on pwviou5 studies and our study, the rnosl coninion breakpoints on uterine leiomvomas aw I “q 1:L 1.5, iated hreak1-lq ‘,L’-2.4. anti 7q22- 32. Onc.ogeIle-assc,( points in malignanf and benign tumors c)lfrr exiting opportunities of studying these breakpoints at a moles.ular level. Oncogenes Int 1 and c-j0.s arc 10~atetl in the 1L’cjl ZS-15 and llq2P-L’G regions, respect i\,cly. Consistent real-t‘an~:enleIits in region 12~11S- 15 arc found in a number of tumors, such as liponlas.’ nlwoid liposar(‘o1llas, IT. Ix pleiomorphic adenomas ()C sa’liwry glmd,” ad uterine leic,Invomas.“~‘~“.’ ‘~’’ Two n~olccular studies and uterine IcGonr\ OIIIX~” 0Ii niysoid lil”)sar~oIrias’” hwe shown that oncogeIle INT 1 is not disrlli>tcd bz ~tl~~oI~losorn~11 rearrangements. .A breakpoint in the 1+L’Z-2G rcqion was the most frequelltlv li)und hreakpoint in our study. Whether (.-/o,v in 1-~;~I?!?-!?~ is rearranged bv cranslocation in urerilw It~ion~~oma~ is not known. II is interesting to note that chromosome 1S was broken at the $4 region and the distal portion was translocated to chromosome 1 at the p%i r-rgion in case no. .5. Whether the Kb gene \vas affwred and heterozvgosity was lost bv this translocation is unknown. Breakpoint 1p% was’also found in five unwlated uterine leiomyonxis 1,~ Mark et al. ‘I’ Delerioll :Sp I Yp 14 was found in cases no. ti and 1 1 as a collsislent c~hange. This deletion involving a possible suppressor gene is ver) :. flwlucntly found in malignant tumors. such as sn~all cell carcinoma of the lung,” renal cell carcinonx~.L)’ and carcinema of the uterine cervix.” This mav indicate that loss of tumor suppressor gene may also (Air in benign (Lroup 5:

-1D.4

--

3p13 --

WB

. m \

3~12

l!l m3

I4-

7q22

m4

,12q13 \

12

“lq22

m5

* ?

14

1cq22

---C 12

12q13

!I 14

m6

FIGURE 4. ideograms of marker chromosomes in case no. 6: ml = der(2)t(2;4)(pl3:p16); m2 = der(4)t(2;4)(pl3;pl6): m3 del(3)(p13p14), m4 = del(7)(q22); m5 = der(12)t(12:14) (q13:q22): m6 = der(14)t(12;14)(q13;q22).

1013

(3ianges

HUMAN PATHOLOGY TABLE 1.

Chromosomal

Review of Cytogenetic

Volume 22, No. 10 (October 1991) Findings in Present and Previous Studies No. ot CaSeS

Reference

Changes

Group 1 t(12;14)(q14-15;q23-24)

I 1 2 3 4 1” 2 1* 1* 2* 3* 1 2* 3* 4 5 6 1 1 1*

5 7

11 12 Pre 14

5 5 6 12 Pre

dir ins(12;14)(pll;q22q24) dir ins(14;12)(q22-23;ql4-15q23-24) t(12;22)(q15:q13),del(l4)(qll.2q23-24) t[inv(l2)(915q23);14] t[inv(l2)(p13q15);14][pl3;q24] t(12;13;14)(q15;q31;q22) Group 2 t(12;22)(q15;q13) t(X;l?)(p22.3;q15) dir ins(l:12)(q21:p13q24.3) t(4;12)(ql2;plI.2) t(2;12)(q37;q14) 1(2;12)(q13;q15) inv(12)(q15q24)

12 13 12

Group 3 t(6;14)(q23:p23-~25) t(2;14)(pll;pll) t(l‘I;?)(q?S-24;‘) r(X;5;14)(q24?;p15;q24) t(X;3;14)(q26;q27;q22) t(14;15)(q12?p12?)

8 7 Pre

Group 4 del(7)

14 10 I? Pre

Group 5 t(1;2)(p36;p24)

12

Croup 6 del(S)(pl3pl4) ins(1;4)(q32;q31q35) t(l;ti)(q23;p2 1) t(6:13)(p23;q?l) t(4;11)(q13;q25) inv(4)(p15q12) invW(p25q2 1) inv(3)(pl?q24) del(lS)(qllq22) del(lO)(q22q24)

P1-e

15

Group 7 Trisomy 7 Trisomy 12

Pre 14

Group 8 Normal karyotypes Abbreviation: Pre. present study. * Cases with additional chromosomal t New cases.

abnormalities.

1014

7/7

l/Y 4/34

l/15 3/39

1 2

7/10 no/so 37/37 59/84 24/25 2X/28

l/9 l/9 l/5 l/l5 l/39 I /39

1* 1 1 1* 1* 1* 1

37/37 22/26 so/30 29/29 u/44 0.60 (i/39

‘/5 l/21 l/21 l/?l l/15 l/IS l/15

1 I 1 2 Y 4

SO/46 2/4 YO/30 4/24 27/27 6/20

l/’

1 1 i’ 1 1 ? 3

I”/13 4/25 6/sO 7/58 ‘4/20 4/w 5/“3

I /20 2/?

1 ” 3 1 :i

1O/40 44/M 15/15

5/15

I 2 1 2 :1* 3* 5 ti 7*

14

No. of Abnormal Cases/Total Cases

Cells/Total No. of Cells Studied

9/13 3/4 Yjl3 15/15 10/l 1 35/42 33/33 25/z 32/32 ‘Lo/20 5/5 lO/lO 17/17 3/-L

1

6 9 9 9

No. of Abnormal

6/24f

l/34 l/39 l/39 l/39 l/39

l/15 s/39

It'/12 60/60 4/N

1/w

I

-I/l6 S/9 S/8 1 l/l 1 4/5 I_(/15 3/!, X/S 12/20

I /:w 1/13t 1/13t I/13t 1/13t 1/13t 1/13t 1/13t ‘/l

I I

Y/20 l4/31

l/39 1/31t

CYTOGENETICS

OF UTERINE LEIOMYOMAS

(Hu & Surti)

6

FIGURE 5. Chromosome breakpoints observed in 39 cases. ‘1, Observed in three or more cells; D, observed in one or two cells.

10

1

k

D

13

16

16

l

8

19

leiomyomas. Monosomy 22, founcl in cases no. 2 and 4, has been frequently found in meningiomas and was noted by Gibas et al in one of five cases of uterine leiomyomas.” It has also been observed in retroperitoneal, peritoneal, intestinal, and stomach leiomyosarconlas,‘“-“i Finding monosomy 22 in benign uterine leiomyomas suggests a similar cytogenetic mechanism in benign an’d malignant proliferation of smooth muscle cells. In case no. 15, we analyzed 20 cells and found three with tnsomy 7. Chromosome 7 has been associated with enhanc’ed metastatic potential and invasiveness by overexpression of epidermal growth factor in somatic cell hybrids”* and bladder cancer.‘” Trisomy 7 has been found in several different malignant solid tumors, such as glioblastoma”” and bladder cancer.” It would be of interest to see if trisomy 7 also occurs in uterine leiomyosarcomas. Some of the single-cell structural abnormalities in cases with normal karyotype were the same as the ones fSound as consistent changes in 15 abnormal cases, such as csb( 12)(ql5) in one cell each (four cases), ctb tr(l1; 1 S!)(q23;q15) in one cell (one case), and ctb( 13)(q14) in one cell (one case). Based on these data, it is possible that cytogenetic changes may represent secondary change after the occurrence of a primary event that may promote genetic instability. The nonrandom nature of the chromosomal changes indicates uterine

21

20

8

22

the presence of genes involved in the formation of leiomyoma at the breakpoints observed, and can aid in investigating the responsible molecular events. The eight cytogenetic subgroups did not seem to be associated with specific histologic features in this small study. All cases were typical leiomyomas. with the exception of two cases: one case (no. 10) with a histology of a bizarre leiomyoma and another case of cellular leiomyoma with a normal karyotype. More cases with different histologic features need to be analyzed cytogenetically before firm conclusions can be made. :~rknourlrrlgrn(J,lt.

Leger &rig0

The authors would like IO thank William 1,ori Hoffner for technical assistanc.c. and Sail) fol- typing the manuscript.

and

REFERENCES

1015

HUMAN PATHOLOGY

Volume 22, No. 10 (October

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18. Mandahl N. Hrim S, Kennet K, vt al: C:lonal chromosome abnormalities in two liposarcomas. Cancer Genet C:ytogenrt 28: 137l-14. 1987 19. Turc-Care1 C:, Pietrlak E, Kakati S, rt al: The human INTI gene is locared at chr~omosome r-egion I2q 1% 12q 13 and is not rearranged in myxoid liposarwma with t( 12: 16)(q 13: p I 1). On< ogcne Res 1:397-405, 1987 20. Arheden Ii, Nilbert M. Heim S. et al: No amplification 01 rearrangement of INTl. GI.1. or CX)L’LAl in uterine leiomyomas with t(l2; l~)(ql-1-15;q23-24). C:ancer-Genrr Cytogenet 39:195-201. 1989 2 I. Whang-peng J, Kw-shan CS, Lw EC.: Specific chromosome defect associated with human small cell lung cancer: 3P( 11-23).Scienw 215:181-182, 19x2 2?. Kovacs G. Erlandsson R. Boldog F: C:onsistcnt chromosome 3p deletion and loss of hrtcrwygosiry in renal cell cat.cinom;l. Proc Nat1 Acad Sci I_?%48X:1571-1575. 1988 25. Yokota J, Tsukada Y. Nakajima T, et al: Los?, of hrttw)rygosit\ on rhe shol-t arm of chromosome :iin carcinoma of the ulerinc cervix. Cancel- Res 49:3598-360 1. 11189 21. Mark J: G-band analysis of a human intestinal leiomyosarcoma. Acta Pathol Microbial Stand X4:538-510. 1976 25. Dal
1016

19Xli