Cytogenetic and Fluorescence In Situ Hybridization Analysis of Breast Fibroadenomas Christine F. Stephenson, Roy I. Davis, George E. Moore, and Avery A. Sandberg
ABSTRACT: We report cytogenetic and fluorescence in situ hybridization (FISH) analysis findings in 7 patients with breast fibroadenomas (FA). Three patients were cytogenetically abnormal. One patient had a translocation t(3;5)(p22;q13), the second had trisomy 8, and the third had two clones, 47,XX, ÷ 11 and 47,XX, + 10.
INTRODUCTION Fibroadenomas (FA) are the most common benign solid tumors of the female breast, occurring most frequently in women in their third decade, and may be multiple in 13-20% of patients [1, 2]. FA appear as rubbery-firm, smooth, and excessively mobile lesions [3]. They are characterized by proliferation of both epithelial and mesenchymal elements; the primary neoplastic component of fibroadenomas arising from stromal (mesenchymal) proliferation [4]. Malignant changes of the epithelium component giving rise to carcinoma in situ can occur but are believed to be rare [5]. Cytogenetic abnormalities in primary benign breast tumors have been described; although the literature is extremely limited [4, 6-8]. Consequently, the significance of these changes remains unclear. In this preliminary study, we report cytogenetic abnormalities in three of seven FA. PATIENTS AND METHODS A sample of each breast FA was received for cytogenetic analysis after surgical excision from 7 women, age range 18-49 years. The histological diagnosis in all cases was FA. Cytogenetics Chromosome analysis of the tumor was performed on shortterm cultures as described previously [9]. Fresh tumor sample was collected aseptically during surgery and shipped
From The Cancer Center of Southwest Biomedical Research Institute, Scottsdale (C. F. S., A. A. A.), Department of Pathology, St. Joseph's Hospital and Medical Center, Phoenix (R. I. D.), Arizona, and Division of Surgical Oncology, Department of Health and Hospitals, Denver, Colorado (G. E. M.) Address reprint requests to: Christine F. Stephenson, The Cancer Center of Southwest Biomedical Research Institute, 6401 E. Thomas Road, Scottsdale, AZ 85251. Received February 14, 1992; accepted April 3, 1992. 32 Cancer Genet Cytogenet63:32-36 (1992) 0165-4608/92/$05.00
to our laboratory in sterile medium. The tumor was disaggregated in collagenase (200 U/ml) overnight. The following day, the suspension was seeded in flasks and on coverslips with RPMI 1640 medium supplemented with 17% fetal bovine serum, L-glutamine (200 mM), and penicillinstreptomycin (5,000 U/ml; 5,000 /~g/ml) and placed in a humidified atmosphere with 5% CO2 for a period of 5-7 days. Harvest was performed after overnight Colcemid exposure (0.01 /~g/ml), hypotonic shock (0.48% HEPES, 0.02% EGTA, 0.3% KCI), and fixation with methanol : acetic acid (3:1). Air-dried chromosome preparations were Gbanded using trypsin. The karyotypes were expressed according to the International System for Human Cytogenetic Nomenclature [10]. Fluorescence In Situ Hybridization (FISH) Slides were stored in 70% ethanol at 4°C and air dried immediately before use. Centromeric probes for chromosomes 8,10, and 11 were used. The probes for chromosomes 10 and 11 were purchased in a biotin-labeled form from Oncor. The probe for chromosome 8 was obtained from D. T. Donlon, Stanford University Medical Center [11], and was labeled as follows. Plasmid DNA was nicked using DNAase and then labeled with biotin-11-dUTP (Enzo Diagnostics) by the random primer method of Feinberg and Vogelstein [12]. Hybridization and detection were performed according to the method of Pinkel et al. [13] with modifications. Slides were denatured for 2 minutes at 70°C in 70% formamide, 2 x SSC, pH 7.0, dehydrated in a cold ethanol series, and air dried. A hybridization mixture of 55% formamide, 10% dextran sulfate, 1 x SSC, 50/~g/ml carrier DNA, and 100 ng/ml probe. DNA was denatured for 5 minutes at 70°C and applied to a prewarmed slide which was then incubated overnight at 37°C, followed by room temperature washes in 2 x SSC, pH 7.0, and PN buffer (0.1 M NaPO4, pH 8.0; 0.1% NP-40). Fluoresceinated avidin (Vector Laboratories), [5 /~g/ml in PNM buffer (PN buffer with 5% non-fat dry milk, 0.2% NAN2)] was applied, followed by biotinylated antiavidin (Vector) 5/.~g/ml in PNM © 1992 Elsevier SciencePublishing Co., Inc. 655 Avenueof the Americas, New York, NY 10010
Cytogenetic and FISH Analysis of F A
Table 1
33
Cytogenetic findings in 7 patients with breast fibroadenoma
Patient
Laboratory no.
Age (yr)
Diagnosis
Karyotype (no. of cells)
1
ST-91-07853
42
2
ST-91-07855
29
FA Lipoma FA
3 4 5 6 7
ST-91-17485 ST-91-30750 ST-91-32075 ST-91-35384 ST-91-38519
20 30 49 38 18
FA FA FA FA FA
46,XX,t(3;5)(p22;q13)[20] 46,XX[20] 46,XX[15] 47,XX, + 811] 46,XX[22] 46,XX[20] 46,XX[20] 46,XX[20] 47,XX, + 1114]/47,XX, + 1011]/46,XX[15]
buffer and a second layer of fluoresceinated avidin. Coverslips were m o u n t e d w i t h an antifade solution of p-phenyo l e n e d i a m i n e d i h y d r o c h l o r i d e containing p r o p i d i u m iodide (1 ~g/ml).
RESULTS The cytogenetic data are presented in detail in Table 1. Three of the seven tumors had cytogenetic abnormalities. Case 1 exhibited a translocation t(3;5)(p22;q13) in all cells analyzed (Fig. 1). This is not a constitutional variant since
Figure 1
cells analyzed from a l i p o m a from the same patient were normal (i.e., 46,XX) and d i d not have the translocation. Case 7 had a clone containing +11 (Fig. 2) and one cell with + 10 (Fig. 3). FISH d e m o n s t r a t e d that 15% (control 1.7 - 0.7) of the cells were trisomic for c h r o m o s o m e 11 (Fig. 4) and that 2.4% (control 1.0 +- 0.2%) of the cells were positive for + 10 (Fig. 5). Case 2 had one cell that cytogenetically was 47,XX,+ 8. FISH s h o w e d that 2% of the interphase nuclei were + 8 (control 0.4 -+ 0.3). FISH results are shown in detail in Table 2. Controls for probes 8, 10, and 11 were performed using p e r i p h e r a l blood taken
G-banded karyotype from patient 1 showing t(3;5)(p22;q13).
!
1
2
4
8
5
-I 8
D _
M
7'
8
9
10
11
12
t!t -'
_
.....
13 I-----'
s . . . . . 19
14
15
18
:'! .....
18
17
~;f-----| 20
21
22
Y
X
34
_
1
2
_
.
4
8
5
_____~:~ _
~ w
B
7'
8
g
10
11
I-- "~" 13
14
16
12
"~ ~o :~
IB
17
18
- -I
i. . . . . . . . . . . . . .
fg
i. . . .
20
21
~2
I
¥ ×
Figure 2
G-banded karyotype from patient 7 showing + 11.
Figure 3
G-banded karyotype from patient 7 showing + 10.
I
i_
O
2
8
7
B
i_
I~-
9
10
F-.;~al 13 41oi
lg
14
.....
-I
i_
15
16
.....
4
B
11
12
~
.... 1~r
~-~-I 18
v,T.--I
20
21
22
¥
Cytogenetic and FISH A n a l y s i s of FA
35
Figure 4 Interphase nuclei demonstrating two and three signals, respectively, for chromosome 11 in patient 7.
Figure 5 Interphase nucleus demonstrating three signals for chromosome 10 in patient 7.
from hematologically normal patients. For each control, 1,000 interphase nuclei were counted. The remaining four tumors were cytogenetically normal.
observed in benign tumors they are generally accepted as primary neoplasia-associated abnormalities [14]. Fletcher et al. [8] described two F A in w h i c h t r i s o m y 11 was observed as a clone; one case on its o w n (i.e., 47,XX,+11) and the other with a d d i t i o n a l n u m e r i c a l and structural abnormalities. Using i m m u n o h i s t o c h e m i c a l techniques (alkaline phosphatase antialkaline phosphatase, APAAP) and antivimentin, and antikeratin m o n o c l o n a l antibodies, they were able to demonstrate that cells with the trisomy 11 (vimentin-positive) were derived from the m e s e n c h y m a l element, confirming that proliferation of the m e s e n c h y m a l cells represents the neoplastic proliferation in FA. In the biclonal FA reported in our study, the trisomy 11 probably arose in cells of m e s e n c h y m a l origin. The + 10 cells also could have arisen from the m e s e n c h y m a l cells, but m a y have arisen from the epithelial c o m p o n e n t of the tumor. The significance of the biclonal findings is unclear, but the trisomy 10 (if of epithelial origin) m a y be indicative of a premalignant change. Malignant transformation (i.e., dev e l o p m e n t of carcinoma in situ, w h e t h e r lobular or ductal) in FA is rare [5, 15, 16], as are reports of b i c l o n a l i t y in F A (to our knowledge, this is the first). The significance of these findings and their role in the tumorigenesis of F A will become more evident 1) as more samples are studied; 2) with determination of the origin (lineage) of abnormal cells, particularly in biclonal cases; and 3) w i t h close follow-up of w o m e n who have tumors with biclonal and abnormal findings.
DISCUSSION FA represent the most c o m m o n benign tumor of the breast in young w o m e n [2], however literature reports describing c h r o m o s o m e aberrations in these tumors are limited [4, 6-8]. These reports show that generally only approximately 2 0 - 3 0 % of FA are cytogenetically abnormal. No specific changes have been recorded, and no preferential involvement of a c h r o m o s o m e or c h r o m o s o m e region has been observed. A b n o r m a l i t i e s of c h r o m o s o m e 12 have been reported in three cases by Calabrese et al. [6], but the rearrangements were not consistent. In two cases, the breakpoint on 12 is p12; both cases involve translocations, but the c h r o m o s o m e s are different, i.e., t(12;21)(p12;q22) in one case and t(8;12)(q24;p12) in the other. In the third, the breakpoint was q15. The breakpoint 'p12 on chromosome 12 m a y p l a y a role in the tumorigenesis of FA, but at this stage its precise function is unknown. In our study, the abnormalities were different and were observed in three of seven tumors. In one case, a translocation t(3;5)(p22;q13), w h i c h although present in all cells karyotyped, was not constitutional because a l i p o m a analyzed from the same patient was cytogenetically normal. The second abnormal tumor was characterized by trisomy 8. The third case was biclonah 47,XX,+11 (15% of cells) and 47,XX, + 10 (2.4% of cells). Clonal n u m e r i c a l aberrations are a c o m m o n feature in benign and malignant tumors; consequently, when they are
This work was supported in part by Grant No. CA-41183 from the National Cancer Institute.
36
C . F . S t e p h e n s o n et al.
Table 2
FISH r e s u l t s s h o w i n g s i g n a l d i s t r i b u t i o n for i n t e r p h a s e n u c l e i No. of Signals (%)
Case
Chromosome
No. of nuclei scored
0
1
2
3
1.0 0.0
4.0 2.9 ± 0.3
92.0 96.7 ± 0.05
2.0 0.4 ± 0.3
1.0 0
200 1,000
0.8 0.1 ± 0.1
3.2 8.3 ± 0.6
93.8 90.2 ± 0.7
2.4 1.0 ± 0.2
0 0
500 1,000
0.4 0.5 ± 0.6
0.8 8.4 ± 0.6
83.2 89.3 ± 2.0
15.0 1.7 ± 0.7
0.8 0
500 1,000
8
T C 10 T C 11 T C
Abbreviations: T, tumor; C, control values ± SD.
The authors thank Mary Powell for technical assistance, Shirley Frazzini for secretarial expertise, Robert Roeder and Debbie Griggs for photographic assistance, and Edward J. Donahue for his assistance.
REFERENCES
1. Harris JR, Hellman S, Henderson IC, Kinne DW (1987): Breast Disorders, J. B. Lippincott, Philadelphia, p. 22. 2. Willkinson S, Forrest APM (1985): Fibro-adenoma of the breast. Br J Surg 72:838-840. 3. Hughes LE, Mansel RE, Webster DJT (1989): Benign Disorders and Diseases of the Breast: Concepts and Clinical Management. Bailliere Tindall, London. 4. Fletcher JA, Pinkus GS, Morton CC (1989): Combined immunohistochemical/cytogenetic (IH/C) approach reveals lineage specificity of chromosome aberrations: Application to solid tumors (abstract). Am J Hum Genet 45:A21. 5. Diaz NM, Palmer JO, McDivitt RW (1991) Carcinoma arising within fibroadenomas of the breast: A clinicopathologic study of 105 patients. Am J Clin Pathol 95:614-622. 6. Calabrese G, Di Virgilio C, Cianchetti E, Franchi PG, Stuppia L, Parruti G, Bianchi PG, Palka G (1991): Chromosome abnormalities in breast fibroadenomas. Genes Chromosomes Cancer 3:202-204. 7. Khishin AF, A1-Zwahri MM, Hassan AA, Morad MM (1976): Cytogenetic studies of benign and malignant tumors of the
breast. In S Amedares, R Lisker (eds.): International Congress of Human Genetics, abstract 347 pp. 133-134. Mexico, October 10-15, 1976. 8. Fletcher JA, Pinkus GS, Weidner N, Morton CC (1991): Lineage-restricted clonality in biphasic solid tumors. Am J Pathol 138:1199-1207. 9. Limon J, Dal Cin P, Sandberg AA (1986): Application of long term collagenase disaggregation for the cytogenetic analysis of solid tumors. Cancer Genet Cytogenet 23:305-313. 10. ISCN (1991): Guidelines for Cancer Cytogenetics, Supplement to An International System for Human Cytogenetic Nomenclature, F Mitelman (ed.), Karger, Basel, 1991. 11. Donlon T, Wyman AR, Mulholland J, Balker D, Bruns G, Lah S, Bostein D (1986): Alpha satellite-like sequences at the centromere of chromosome 8. Am J Hum Genet 39:A196. 12. Feinberg AP, Vogelstein B (1983): A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 132:6-13. 13. Pinkel D, Straume T, Gray JW (1986): Cytogenetic analysis using quantitative, high-sensitivity, fluorescence hybridization. Proc Natl Acad Sci USA 83:2934-2938. 14. Helm S, Mitelman F (1987): Cancer Cytogenetics. Alan R. Liss, New York. 15. Pick PW, Lossifides IA (1991): Occurrence of breast carcinoma within a fibroadenoma. Arch Pathol Lab Med 108:590594. 16. Gupta RK, Simpson J (1991): Carcinoma of the breast in a fibroadenoma: Diagnosis by finemeedle aspiration cytology. Diag Cytopathol 7:60-62.
ABSTRACT: We report cytogenetic and fluorescence in situ hybridization (FISH) analysis findings in 7 patients with breast fibroadenomas (FA). Three patients were cytogenetically abnormal. One patient had a translocation t(3;5)(p22;q13), the second had trisomy 8, and the third had two clones, 47,XX, ÷ 11 and 47,XX, + 10.
INTRODUCTION Fibroadenomas (FA) are the most common benign solid tumors of the female breast, occurring most frequently in women in their third decade, and may be multiple in 13-20% of patients [1, 2]. FA appear as rubbery-firm, smooth, and excessively mobile lesions [3]. They are characterized by proliferation of both epithelial and mesenchymal elements; the primary neoplastic component of fibroadenomas arising from stromal (mesenchymal) proliferation [4]. Malignant changes of the epithelium component giving rise to carcinoma in situ can occur but are believed to be rare [5]. Cytogenetic abnormalities in primary benign breast tumors have been described; although the literature is extremely limited [4, 6-8]. Consequently, the significance of these changes remains unclear. In this preliminary study, we report cytogenetic abnormalities in three of seven FA. PATIENTS AND METHODS A sample of each breast FA was received for cytogenetic analysis after surgical excision from 7 women, age range 18-49 years. The histological diagnosis in all cases was FA. Cytogenetics Chromosome analysis of the tumor was performed on shortterm cultures as described previously [9]. Fresh tumor sample was collected aseptically during surgery and shipped
From The Cancer Center of Southwest Biomedical Research Institute, Scottsdale (C. F. S., A. A. A.), Department of Pathology, St. Joseph's Hospital and Medical Center, Phoenix (R. I. D.), Arizona, and Division of Surgical Oncology, Department of Health and Hospitals, Denver, Colorado (G. E. M.) Address reprint requests to: Christine F. Stephenson, The Cancer Center of Southwest Biomedical Research Institute, 6401 E. Thomas Road, Scottsdale, AZ 85251. Received February 14, 1992; accepted April 3, 1992. 32 Cancer Genet Cytogenet63:32-36 (1992) 0165-4608/92/$05.00
to our laboratory in sterile medium. The tumor was disaggregated in collagenase (200 U/ml) overnight. The following day, the suspension was seeded in flasks and on coverslips with RPMI 1640 medium supplemented with 17% fetal bovine serum, L-glutamine (200 mM), and penicillinstreptomycin (5,000 U/ml; 5,000 /~g/ml) and placed in a humidified atmosphere with 5% CO2 for a period of 5-7 days. Harvest was performed after overnight Colcemid exposure (0.01 /~g/ml), hypotonic shock (0.48% HEPES, 0.02% EGTA, 0.3% KCI), and fixation with methanol : acetic acid (3:1). Air-dried chromosome preparations were Gbanded using trypsin. The karyotypes were expressed according to the International System for Human Cytogenetic Nomenclature [10]. Fluorescence In Situ Hybridization (FISH) Slides were stored in 70% ethanol at 4°C and air dried immediately before use. Centromeric probes for chromosomes 8,10, and 11 were used. The probes for chromosomes 10 and 11 were purchased in a biotin-labeled form from Oncor. The probe for chromosome 8 was obtained from D. T. Donlon, Stanford University Medical Center [11], and was labeled as follows. Plasmid DNA was nicked using DNAase and then labeled with biotin-11-dUTP (Enzo Diagnostics) by the random primer method of Feinberg and Vogelstein [12]. Hybridization and detection were performed according to the method of Pinkel et al. [13] with modifications. Slides were denatured for 2 minutes at 70°C in 70% formamide, 2 x SSC, pH 7.0, dehydrated in a cold ethanol series, and air dried. A hybridization mixture of 55% formamide, 10% dextran sulfate, 1 x SSC, 50/~g/ml carrier DNA, and 100 ng/ml probe. DNA was denatured for 5 minutes at 70°C and applied to a prewarmed slide which was then incubated overnight at 37°C, followed by room temperature washes in 2 x SSC, pH 7.0, and PN buffer (0.1 M NaPO4, pH 8.0; 0.1% NP-40). Fluoresceinated avidin (Vector Laboratories), [5 /~g/ml in PNM buffer (PN buffer with 5% non-fat dry milk, 0.2% NAN2)] was applied, followed by biotinylated antiavidin (Vector) 5/.~g/ml in PNM © 1992 Elsevier SciencePublishing Co., Inc. 655 Avenueof the Americas, New York, NY 10010
Cytogenetic and FISH Analysis of F A
Table 1
33
Cytogenetic findings in 7 patients with breast fibroadenoma
Patient
Laboratory no.
Age (yr)
Diagnosis
Karyotype (no. of cells)
1
ST-91-07853
42
2
ST-91-07855
29
FA Lipoma FA
3 4 5 6 7
ST-91-17485 ST-91-30750 ST-91-32075 ST-91-35384 ST-91-38519
20 30 49 38 18
FA FA FA FA FA
46,XX,t(3;5)(p22;q13)[20] 46,XX[20] 46,XX[15] 47,XX, + 811] 46,XX[22] 46,XX[20] 46,XX[20] 46,XX[20] 47,XX, + 1114]/47,XX, + 1011]/46,XX[15]
buffer and a second layer of fluoresceinated avidin. Coverslips were m o u n t e d w i t h an antifade solution of p-phenyo l e n e d i a m i n e d i h y d r o c h l o r i d e containing p r o p i d i u m iodide (1 ~g/ml).
RESULTS The cytogenetic data are presented in detail in Table 1. Three of the seven tumors had cytogenetic abnormalities. Case 1 exhibited a translocation t(3;5)(p22;q13) in all cells analyzed (Fig. 1). This is not a constitutional variant since
Figure 1
cells analyzed from a l i p o m a from the same patient were normal (i.e., 46,XX) and d i d not have the translocation. Case 7 had a clone containing +11 (Fig. 2) and one cell with + 10 (Fig. 3). FISH d e m o n s t r a t e d that 15% (control 1.7 - 0.7) of the cells were trisomic for c h r o m o s o m e 11 (Fig. 4) and that 2.4% (control 1.0 +- 0.2%) of the cells were positive for + 10 (Fig. 5). Case 2 had one cell that cytogenetically was 47,XX,+ 8. FISH s h o w e d that 2% of the interphase nuclei were + 8 (control 0.4 -+ 0.3). FISH results are shown in detail in Table 2. Controls for probes 8, 10, and 11 were performed using p e r i p h e r a l blood taken
G-banded karyotype from patient 1 showing t(3;5)(p22;q13).
!
1
2
4
8
5
-I 8
D _
M
7'
8
9
10
11
12
t!t -'
_
.....
13 I-----'
s . . . . . 19
14
15
18
:'! .....
18
17
~;f-----| 20
21
22
Y
X
34
_
1
2
_
.
4
8
5
_____~:~ _
~ w
B
7'
8
g
10
11
I-- "~" 13
14
16
12
"~ ~o :~
IB
17
18
- -I
i. . . . . . . . . . . . . .
fg
i. . . .
20
21
~2
I
¥ ×
Figure 2
G-banded karyotype from patient 7 showing + 11.
Figure 3
G-banded karyotype from patient 7 showing + 10.
I
i_
O
2
8
7
B
i_
I~-
9
10
F-.;~al 13 41oi
lg
14
.....
-I
i_
15
16
.....
4
B
11
12
~
.... 1~r
~-~-I 18
v,T.--I
20
21
22
¥
Cytogenetic and FISH A n a l y s i s of FA
35
Figure 4 Interphase nuclei demonstrating two and three signals, respectively, for chromosome 11 in patient 7.
Figure 5 Interphase nucleus demonstrating three signals for chromosome 10 in patient 7.
from hematologically normal patients. For each control, 1,000 interphase nuclei were counted. The remaining four tumors were cytogenetically normal.
observed in benign tumors they are generally accepted as primary neoplasia-associated abnormalities [14]. Fletcher et al. [8] described two F A in w h i c h t r i s o m y 11 was observed as a clone; one case on its o w n (i.e., 47,XX,+11) and the other with a d d i t i o n a l n u m e r i c a l and structural abnormalities. Using i m m u n o h i s t o c h e m i c a l techniques (alkaline phosphatase antialkaline phosphatase, APAAP) and antivimentin, and antikeratin m o n o c l o n a l antibodies, they were able to demonstrate that cells with the trisomy 11 (vimentin-positive) were derived from the m e s e n c h y m a l element, confirming that proliferation of the m e s e n c h y m a l cells represents the neoplastic proliferation in FA. In the biclonal FA reported in our study, the trisomy 11 probably arose in cells of m e s e n c h y m a l origin. The + 10 cells also could have arisen from the m e s e n c h y m a l cells, but m a y have arisen from the epithelial c o m p o n e n t of the tumor. The significance of the biclonal findings is unclear, but the trisomy 10 (if of epithelial origin) m a y be indicative of a premalignant change. Malignant transformation (i.e., dev e l o p m e n t of carcinoma in situ, w h e t h e r lobular or ductal) in FA is rare [5, 15, 16], as are reports of b i c l o n a l i t y in F A (to our knowledge, this is the first). The significance of these findings and their role in the tumorigenesis of F A will become more evident 1) as more samples are studied; 2) with determination of the origin (lineage) of abnormal cells, particularly in biclonal cases; and 3) w i t h close follow-up of w o m e n who have tumors with biclonal and abnormal findings.
DISCUSSION FA represent the most c o m m o n benign tumor of the breast in young w o m e n [2], however literature reports describing c h r o m o s o m e aberrations in these tumors are limited [4, 6-8]. These reports show that generally only approximately 2 0 - 3 0 % of FA are cytogenetically abnormal. No specific changes have been recorded, and no preferential involvement of a c h r o m o s o m e or c h r o m o s o m e region has been observed. A b n o r m a l i t i e s of c h r o m o s o m e 12 have been reported in three cases by Calabrese et al. [6], but the rearrangements were not consistent. In two cases, the breakpoint on 12 is p12; both cases involve translocations, but the c h r o m o s o m e s are different, i.e., t(12;21)(p12;q22) in one case and t(8;12)(q24;p12) in the other. In the third, the breakpoint was q15. The breakpoint 'p12 on chromosome 12 m a y p l a y a role in the tumorigenesis of FA, but at this stage its precise function is unknown. In our study, the abnormalities were different and were observed in three of seven tumors. In one case, a translocation t(3;5)(p22;q13), w h i c h although present in all cells karyotyped, was not constitutional because a l i p o m a analyzed from the same patient was cytogenetically normal. The second abnormal tumor was characterized by trisomy 8. The third case was biclonah 47,XX,+11 (15% of cells) and 47,XX, + 10 (2.4% of cells). Clonal n u m e r i c a l aberrations are a c o m m o n feature in benign and malignant tumors; consequently, when they are
This work was supported in part by Grant No. CA-41183 from the National Cancer Institute.
36
C . F . S t e p h e n s o n et al.
Table 2
FISH r e s u l t s s h o w i n g s i g n a l d i s t r i b u t i o n for i n t e r p h a s e n u c l e i No. of Signals (%)
Case
Chromosome
No. of nuclei scored
0
1
2
3
1.0 0.0
4.0 2.9 ± 0.3
92.0 96.7 ± 0.05
2.0 0.4 ± 0.3
1.0 0
200 1,000
0.8 0.1 ± 0.1
3.2 8.3 ± 0.6
93.8 90.2 ± 0.7
2.4 1.0 ± 0.2
0 0
500 1,000
0.4 0.5 ± 0.6
0.8 8.4 ± 0.6
83.2 89.3 ± 2.0
15.0 1.7 ± 0.7
0.8 0
500 1,000
8
T C 10 T C 11 T C
Abbreviations: T, tumor; C, control values ± SD.
The authors thank Mary Powell for technical assistance, Shirley Frazzini for secretarial expertise, Robert Roeder and Debbie Griggs for photographic assistance, and Edward J. Donahue for his assistance.
REFERENCES
1. Harris JR, Hellman S, Henderson IC, Kinne DW (1987): Breast Disorders, J. B. Lippincott, Philadelphia, p. 22. 2. Willkinson S, Forrest APM (1985): Fibro-adenoma of the breast. Br J Surg 72:838-840. 3. Hughes LE, Mansel RE, Webster DJT (1989): Benign Disorders and Diseases of the Breast: Concepts and Clinical Management. Bailliere Tindall, London. 4. Fletcher JA, Pinkus GS, Morton CC (1989): Combined immunohistochemical/cytogenetic (IH/C) approach reveals lineage specificity of chromosome aberrations: Application to solid tumors (abstract). Am J Hum Genet 45:A21. 5. Diaz NM, Palmer JO, McDivitt RW (1991) Carcinoma arising within fibroadenomas of the breast: A clinicopathologic study of 105 patients. Am J Clin Pathol 95:614-622. 6. Calabrese G, Di Virgilio C, Cianchetti E, Franchi PG, Stuppia L, Parruti G, Bianchi PG, Palka G (1991): Chromosome abnormalities in breast fibroadenomas. Genes Chromosomes Cancer 3:202-204. 7. Khishin AF, A1-Zwahri MM, Hassan AA, Morad MM (1976): Cytogenetic studies of benign and malignant tumors of the
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