ductal carcinoma in situ: overview and updates
DESCRIPTION
Ductal carcinoma in situ (DCIS) refers to breast epithelial cells that have become “cancerous” but still reside in their normal place in the ducts and lobules. In this setting, cancerous means that there is an abnormal increase in the growth of the epithelial cells, which accumulate within and greatly expand the ducts and lobules (Figure 1). DCIS is a non-lethal type of cancer because it stays in its normal place. However, DCIS is very important because it is the immediate precursor of invasive breast cancers (IBCs), which are potentially lethal. This presentation will provide a general overview of DCIS, including historical perspective, and methods of classification, as well as an update regarding recent important advances in understanding the development and progression of DCIS at the molecular level.TRANSCRIPT
Ductal Carcinoma In Situ (DCIS)of the Human Breast
Overview and Updates
D. Craig Allred, MD
NormalTDLU
EarlyExpansion of
TDLU by DCIS
CompleteExpansion of
TDLU by DCIS
Look Like Ducts “Ductal” Carcinoma In Situ
Definition of DCISBreast epithelial cells that have become "cancerous" but still reside
in their normal place inside the ducts and lobules.
IBC
TDLU
ADHCCHDCIS
ALH LCIS
Time (decades in most cases)
Immediate precursor of invasive breast cancer
Importance of DCIS
TDLU
ADHCCHDCIS
ALH LCIS
Time (decades in most cases)
Late stage in breast cancer evolution
DCIS is the Immediate Precursor of Invasive Breast Cancer (IBC)
Historical Perspective of Invasive Breast Cancer
Ancient
Greece
DarkAges
MiddleAges
Renaissance
Today
Today
1600 BCEdwin Smith
PapyrusEgypt
Historical Perspective of DCIS
Early 1900sFirst Realized DCIS ≠
IBC2-3% all Breast Cancer
Tx = Radical Mastectomy (Cure)
Major Milestones Since:
· DCIS = Precursor of IBCNeeds Detection and Therapy
Screening MammographyDCIS = 20-30% all Breast Cancer
Lumpectomy + Adjuvant Rad ± Tam
Radical Mastectomy
Natural History of DCIS
DCIS(Non-Lethal)
IBC(Potentially Lethal)
TDLU
ADHCCHDCIS
IBC
ALH LCIS
Time (decades in most cases)
TDLU
ADHCCHDCIS
IBC
ALH LCIS
Time (decades in most cases)
Overall Proportion Unknown
≥ 30-40% over 30 yearsJAMA 239:1863, 1978Cancer 46:919, 1980Cancer 49:751, 1982
Sem Diag Pathol 11:223, 1994
Absolute Proportion Too High
All IBCs from undetected DCIS
Classification of DCIS
Comedo Cribriform Solid
PapillaryMicropapillary
Subtypes of DCIS based on gross appearance and predominant microscopic growth pattern
Classification of DCISSubtypes of DCIS based on gross appearance and predominant microscopic growth pattern
Problem = Intra-Tumor Diversity
Solid/Comedo
Cribriform
≥50% DCIS with >1 pattern
Clin Cancer Res 14:339, 2008
Classification of DCIS
Histological Grade:The Degree that Tumor
Resembles Normal
Histological Scoring and Grading of DCIS
Based on Scarff-Bloom-Richardson method of grading IBCClin Cancer Res 14:339, 2008
210 - 75%
intermediate1 - 2
10 - 50%
Score
1> 75%
low< 1
< 10%
3< 10%high> 2
> 50%
Microscopic FeatureA. Glands/papillaeB. Nuclear gradeC. Mitotic rateD. Central necrosis
S= total score (range 4-12)Grade 1 = 4 -7 points (well differentiated)Grade 2 = 8-9 points (intermediate differentiated)Grade 3 = 10-12 points (poorly differentiated)
A=1B=1C=1D=1Total=4Grade =1
A=2B=2C=2D=2Total=8Grade =2
A=3B=3C=3D=3Total=12Grade =3
Classification of DCIS
Histological Grade:The Degree that Tumor
Resembles Normal
Problem = Intra-Tumor Diversity
Grade 3
Grade 1≥50% DCIS
with >1 GradeClin Cancer Res 14:339, 2008
Classification of DCISBiological Features
Standard Prognostic Biomarkers
Histological Score (Modified SBR; Clin Cancer Res 14:339, 2008)
45
67
89
1011
120
Perc
ent
102030405060708090
100
N=200% ER+% PgR+
% p53+% HER2+
% Cases
Pure DCIS
avg %Ki67
WellDifferentiated
PoorlyDifferentiated
45
67
89
1011
12
N=200
56
78
9
N=200
IBC DCIS (+IBC)
34
DCIS IBC
From: Solin, et al: J Clin Oncol 14:754, 1996.
Histological Grade and Standard Biomarkers in DCIS are Related to Rate but not Fate of Progressing to IBC.
LuminalER/PR+GATA3+ERBB2-CK18+Other...
BasalERBB2-ER/PR-CK5+Other…
Mixed
ERBB2ERBB2+ER/PR-GRB7+Other...
Sorlie et al. PNAS 98:10869, 2001
DCIS IBC
Classification of DCISBiological Features
Intrinsic Molecular Subtypes
ERBB2Basal
MixedLuminal
DCISAllred et al. Clin Cancer Res 14:339, 2008
IBC
Grade 1 Grade 2 Grade 3
Ca
se #
020
Ca
se #
012
29.2% 22.5% 2.5%30.0%
9.2%6.6%
30% 60% 10%
10% 85% 5%
Diversity of Histological (Nuclear) Grade Within Cases of DCIS (n=120)
Allred et al. Clin Cancer Res 2008
Category1. No Diversity2. Grade3. Grade + Biol
% Cases51.8%48.2%13.4%
Diversity of Histological Grade and Biological Characteristics Within Cases (n=112)(ER, GATA3, Her2, CK5, CK18 and p53)
(Allred et al. Clin Cancer Res, 2008)
(1 vs. 2 vs. 3)
p=0.0016
% p53+13.8%40.5%45.5%
Exa
mpl
e: C
ase
10
6
H&E
H&E ER=7/8
ER=5/8
Her2=0/8
Her2=6/8 p53=6/8
p53=0/8
Grade 1(60%)
Grade 2(40%)
p=0.2833
% ER+70.7%60.8%57.6%
p=0.8296
% Her2+39.7%39.2%57.6%
Diversity of Histological Grade and Intrinsic
Subtypes Within Cases of DCIS (n=112)(Allred et al. Clin Cancer Res, 2008)
Luminal ALuminal B
Basal
ERBB2
Different SubtypesSame Subtype
73% (11/15) of DCIS with diversity of histological (nuclear) grade and biomarkers
also showed diversity of intrinsic subtypes of nearly all possible combinations
LumA + erbB2 = 4 casesLumA + Basal = 1 caseLumA + LumB = 3 casesLumB + erbB2 = 2 casesBasal + erbB2 = 1 case
Cancer Stem Cell Conundrum
DCIS(Non-Lethal)
IBC(Potentially Lethal)
TDLU
ADHCCHDCIS
IBC
ALH LCIS
Time (decades in most cases)
TDLU
ADHCCHDCIS
IBC
ALH LCIS
Time (decades in most cases)
DCIS ≠ IBC Regarding Invasion
Tumor Epithelial Cells Very SimilarMolec Cancer Res 1:362, 2003
PNAS 100:5974, 2003
Stromal Cells Very Different
Cancer Cell 6:17, 2004
Genetic Differences DCIS vs. IBC
July, 2012
5 Cohorts DCIS vs. IBC54 samples each type50% LCM microdissectedAffymetrix microarrays
Supervised Comparisons≥ 2 Fold; p < 0.05
472 Genes ≥ 1 Group This Study74 Genes ≥ 2 Groups This Study
Meta-analysis:69% Overlap with ≥1 of
all Previous Microarry StudiesAverage = 3.8 Studies/Gene
Range = 3 to 7 Studies
IBC > DCIS (n=42)Candidate Promotersof Tumor Progression
DCIS > IBC (n=32)Candidate Suppressorsof Tumor Progression
Supervised Comparisons≥ 2 Fold; p < 0.05
472 Genes ≥1 Group This Study74 Genes ≥2 Groups This Study
Meta-analysis:38% Overlap with ≥1 of
all Previous Microarry StudiesAverage = 3.8 Studies/Gene
Range = 3 to 5 Studies
Gene Expression Signature (n=74)DCIS vs. IBC
96% Correct Classification Study Samples
Hierarchical Clustering74-Gene Signature
(avg = 34/74 per group)
DCISIBC
Gene Expression Signature (n=74)DCIS vs. IBC
95% Correct Classification Samples in 3 Independent Studies
DCISIBC
Ontologies and Pathways Associated with the Progression of DCIS to IBC
Adjusted p-value
Ontologies and Pathways in Tumor Epithelium vs.Stroma
EMT+
Functional StudiesMammary Intraductal DCIS (MIND) Xenografts
(Behbod et al. Breast Cancer Res, 2009)
DCIS
IBC
DCIS.COM shCSTA-A9
DCIS.COM shCSTA-A9
Functional StudiesHuman DCIS Cell Lines
Cell Line1. DCIS.COM2. DCIS.SUM.2253. DCIS.FSK-H74. hDCIS.015. hDCIS.026. hDCIS.037. hDCIS.048. hTDLU.019. hCCH.0110.hCCH.02
Max Pass #
>100>100
55>100
117
16131510
Keratin IHC+++++
++++++++++++++++++++++++
MIND Xenografts ≤10 WksNone
10%30%30%20%ndndndndndnd
DCIS60%70%70%60%ndndndndndnd
IBC30%0%0%
20%ndndndndndnd
1 2 3 4 5
6 7 8 9 10
Knock Down (shRNAi) “Suppressors”Predict Increased Tumor Progression
1. I-A7 (hairpin)2. I-A9 (hairpin)3. I-D4 (hairpin)4. RFP-1 (control)5. RFP-2 (control)6. DCIS.COM-CBRz (control)
1 2 3 4 5 6
b-Actin
CSTA
Western Blot
3 of 3 hairpinssignificantly decrease protein
expression in DCIS.COM(1A7>IA9>1D4)
Real-time PCR
3 of 3 hairpinssignificantly decrease RNA
expression in DCIS.COM(1A9>1A7>1D4)
complete knockdown
DCIS.COM Transduced with CSTA shRNAi (n=3)
Effects of Knocking
Down “Suppressors
” In Vivo(DCIS.COM)
Normally Suppress Invasion:
CSTA (Cystatin A)DST (Dystonin)
FAT1 (Procadherin)TMEM45A (Transmembrane
Protein 45A)
Validation in Additional DCIS Cell Lines
NormallySuppress Invasion:
CSTADSTFAT1
Validation of CSTA at Protein Levelby IHC in Human Breast Tissue
DCIS
A B
F
C D
E
TDLU
DCIS
DCIS
IBC
IBC
CCH
TDLU
TDLU CCH DCIS IBCCSTA Pos 96% 95% 58% 26%
#Cases 27 40 92 228
P <0.001
Normally Expressedin Myoepithelial Cells
Frequently Mislocalizedto Luminal Cells in DCIS
Significantly Reducedin IBC vs. DCIS
DCIS.COM.shCSTA.1A7 MIND Xenografts
IHC for p63(human and mouse specific)
IHC for Vimentin(human specific)
Human DCIS Cell Lines are Pluripotential(Myoepithelial and Luminal)
MECs
MECs
LUMLUM
pB-pu CILP-pu FAP-pu0.0
20.0
40.0
60.0
80.0
100.0
120.0
> 205 - 200 - 5none
DCIS.COM
Promote Invasion:CILP (Cartilage Intermediate Filament
Protein)FAP (Fibroblast Activation Protein Alpha)
Overexpress (cDNA) “Promoters”Predict Increased Tumor Progression
Improved detection, diagnosis, and surgery (lumpectomy) of DCIS through targeted imaging based on molecular features specific for DCIS vs. IBC.
By understanding more about DCIS we can significantly decrease the incidence of IBC (at least 10%) during the next decade by:
Improved prognostication, therapy, and prevention of DCIS based on targeting molecular features regulating the progression DCIS to IBC.