Introduction
Materials and methods
Study approval and patient samples
DNA and RNA isolation
Comparative genome hybridization based on microarrays
Copy number validation by real-time quantitative PCR
Microarray gene expression analysis
Results
Clinical variable | Category | Number (%) |
---|---|---|
LVIa
| Positive | 25 (43.9) |
Negative | 32 (56.1) | |
SBRb
| I | 9 (15.8) |
II | 17 (29.8) | |
III | 31 (54.4) | |
Clinical stage | I | 9 (15.8) |
II | 21 (36.9) | |
III | 26 (45.6) | |
IV | 1 (1.7) | |
Axillary lymph node | N0 | 20 (35.1) |
N1 | 19 (33.3) | |
N2 | 10 (17.5) | |
N3 | 8 (14.0) | |
Estrogen receptor | Positive | 33 (57.9) |
Negative | 24 (42.1) | |
ERBB2
| Positive | 5 (8.8) |
Negative | 52 (91.2) | |
Triple-negative | Yes | 12 (21.0) |
No | 45 (79.0) |
Total CNAs and their distribution and type according to histological grade
Group | Total events (median) | Gains (median) | Losses (median) | High copy gains (mean) | Homozygous losses (mean) | Mean size (Mb) (median) |
---|---|---|---|---|---|---|
Breast carcinomas (n = 57) | 2,857 (38.0) | 1,477 (23.0) | 1,245 (16.0) | 122 (2.1) | 13 (0.2) | 70.2 (27.3) |
SBR I (n = 9) | 290 (21.0) | 128 (12.0) | 155 (9.0) | 7 (0.8) | 0 (0) | 10.0 (27.0) |
SBR II (n = 17) | 606 (20.0) | 315 (10.0) | 241 (9.0) | 44 (2.6) | 6 (0.3) | 16.2 (26.6) |
SBR III (n = 31) | 1,961* (60.0) | 1,034* (33.0) | 849* (25.0) | 71 (2.3) | 7 (0.2) | 48.9* (27.3) |
LVI positive (n = 25) | 1,289 (38.0) | 676 (25.0) | 558 (14.0) | 51 (2.0) | 4 (0.2) | 33.1 (26.6) |
LVI negative (n = 32) | 1,568 (41.5) | 801 (22.5) | 687 (17.5) | 71 (2.2) | 9 (0.3) | 40.0 (27.6) |
Total CNAs and their distribution and type according to LVI status
Small-scale rearrangements: genomic amplifications and homozygous losses
Known amplicon | Major gene | Total frequency (n affected) | Frequency among histologic grade III samples | Frequency among lymphovascular invasion positive samples |
---|---|---|---|---|
8q24 |
MYC
| 38.6 % (22) |
81.8 %
| 45.4 % |
8p12 |
ZNF703/FGR1
| 31.6 % (18) |
72.2 %
| 44.4 |
8p12 |
FGFR1
| 29.8 % (17) |
64.7 %
| 47.1 % |
19q12 |
CCNE1
| 17.5 % (10) |
100 %
|
55.5 %
|
20q13.2 |
ZNF217
| 14.0 % (8) |
62.5 %
|
62.5 %
|
11q13 |
CCND1
| 14.0 % (8) |
50.0 %
| 25.0 % |
Copy number status of previously reported breast cancer driver genes
Gene expression profiling
Histologic grade IIIa
| Lymphovascular invasion positiveb
| ||||||
---|---|---|---|---|---|---|---|
Gene | Cytoband | Fold-change |
p value | Gene | Cytoband | Fold-change |
p value |
S100A8
| 1q21.3 | 22.0278 | 0.001 |
C19orf33
| 19q13.2 | 4.4608 | 0.021 |
DEFB1
| 8p23.1 | 14.2762 | 0.001 |
LGALS7B
| 19q13.2 | 3.5713 | 0.007 |
ADM
| 11p15.4 | 7.0605 | 0.001 |
CPE
| 4q32.3 | 3.0436 | 0.011 |
MMP1
| 11q22.2 | 6.7464 | 0.001 |
AGBL2
| 11p11.2 | 2.9353 | 0.028 |
MCM10
| 10p13 | 6.4085 | 0.001 |
ARSG
| 17q24.2 | 2.7475 | 0.044 |
TMSB15A
| Xq22.1 | 6.2266 | 0.001 |
UMOD
| 16p12.3 | 2.6808 | 0.025 |
SOX11
| 2p25.2 | 5.5269 | 0.001 |
C13orf31
| 13q14.11 | 2.6631 | 0.026 |
YBX2
| 17p13.1 | 5.3408 | 0.001 |
MYCBPAP
| 17q21.33 | 2.5776 | 0.032 |
KRT81
| 12q13.13 | 5.3214 | 0.001 |
CXXC4
| 4q24 | 2.5767 | 0.044 |
E2F8
| 11p15.1 | 5.2362 | 0.001 |
MED1
| 17q12 | 2.5187 | 0.033 |
NANOS1
| 10q26.11 | 5.1092 | 0.001 |
SHISA5
| 3p21.31 | 2.5084 | 0.002 |
CDC45L
| 22q11.21 | 5.0596 | 0.001 |
GFRA1
| 10q25.3 | −4.4772 | 0.021 |
GRP
| 18q21.32 | −8.9074 | 0.001 |
MFI2
| 3q29 | −4.0174 | 0.029 |
CYP4X1
| 1p33 | −8.1935 | 0.001 |
TBX21
| 17q21.32 | −3.8633 | 0.038 |
SUSD3
| 9q22.31 | −8.0443 | 0.001 |
KRT15
| 17q21.2 | −3.6634 | 0.016 |
ABAT
| 16p13.2 | −7.3311 | 0.001 |
NFASC
| 1q32.1 | −3.3951 | 0.022 |
EVL
| 14q32.2 | −6.9978 | 0.001 |
FZD5
| 2q33.3 | −3.1834 | 0.008 |
KLHDC9
| 1q23.3 | −6.9593 | 0.001 |
TCF7L1
| 2p11.2 | −3.0653 | 0.047 |
C6orf211
| 6q25.1 | −6.9305 | 0.001 |
MYBPC2
| 19q13.33 | −2.8301 | 0.021 |
NEFH
| 22q12.2 | −6.3975 | 0.001 |
CRABP1
| 15q25.1 | −2.7727 | 0.017 |
CX3CR1
| 3p22.2 | −5.3859 | 0.001 |
CIB2
| 15q25.1 | −2.5553 | 0.018 |
SMOC2
| 6q27 | −5.3324 | 0.001 |
DUSP3
| 17q21.31 | −2.5391 | 0.001 |
BTG2
| 1q32.1 | −5.3206 | 0.001 | ||||
FAM198B
| 4q32.1 | −5.0859 | 0.001 | ||||
CCDC74B
| 2q21.1 | −5.0592 | 0.001 |
Summary of the relevant genomic regions and selected genes
Relevant gene(s) | Chromosomal alteration | Clinical association |
---|---|---|
MYC
| 8q24 gain | Preferentially detected in grade III samples |
MTAP
| 9p21 loss | Preferentially detected in grade III samples |
COX6C
| 8q22 gain | Preferentially detected in grade III samples |
PMS2
| 7p22 loss | Preferentially detected in grade III samples |
ERCC8, XRCC4, FBXL17, MEF2C
| 5q11q13 loss | Preferentially detected in grade III samples |
LMO3
| 12p12.3 loss | Preferentially detected in grade III samples |
CCNE1, LOC284395, VSTM2B, POP4, PLEKHF1, C19orf12
| 19q12 minimum common region of gain | Exclusively detected in grade III negative for estrogen-receptor samples |
ADAMTS3
| 4q13.3 amplification | Detected in a grade III LVI-positive sample |
HSD17B12
| 11p11.2 amplification | Detected in a grade III LVI-positive sample |
RERGL, PIK3C2G
| 12p12.3 homozygous deletion | Detected in a grade III LVI-positive sample |
ARHGAP32
| 11q24.3 loss | Preferentially detected in LVI-positive samples |
S100A8, MCM10
| – | Concordant pattern of upregulated gene expression and copy number gains in grade III samples |
GRP, CX3CR1, FAM198B
| – | Concordant pattern of downregulated gene expression and copy number losses in grade III samples |
UMOD, ARSG, MYCBPAP, MED1
| – | Concordant pattern of upregulated gene expression and copy number gains/high gains in LVI-positive samples |
Several
| 16p gain | Preferentially detected in both low-grade and LVI-negative samples (exclusively estrogen-receptor-positive tumors) |