Pathology Journal Reading

Presented by Intern
Objective
To identify the role of cytokeratins
in distinguishing intraductal
papilloma from papillary ductal
carcinoma in situ
Introduction
Papillary breast tumors:
Proliferated mammary epithelium
projects into duct lumen
Intraductal papilloma
Preinvasive papillary ductal carcinoma in
situ (DCIS)

Sometimes difficult to distinguish:
overlapping microscopic appearances
The need to separate them
Distinct biologic behavior
Papillary DCIS potentially progressing
to invasive cancer
Surgical extirpation

Benign papillomas
Discharged from clinical follow up
Cytokeratins (CK)
Fundamental markers of epithelial
differentiation
Cell type & differentiation status

Previous studies:
CK5/CK6, 34BE12 and CK14 in distinguishing
usual epithelial hyperplasia from atypical
ductal hyperplasia (ADH) and DCIS

Presence of myoepithelial cells
Preserved in benign papillomas, scant in papillary
DCIS
Muscle actin and 34BE12 (less specific)


In this study
Detail the expression of three CK
antibody preparation
CK5/6
CK14
34BE12 (recognizing CKs 1, 5, 10 and
14)


Aim of this study
determining their role (CKs) in differentiating the
benign papilloma from malignant in situ papillary
carcinoma

Scant literature that specifically addresses
papillary lesions
previous work focusing on proliferative breast lesions
as a generic group versus DCIS

Semiquantitative criteria of immunoscores to
evaluate the CKs

Ascertain the findings by extrapolating to a
separate group




Materials and Methods
Patients and tumors
50 excision biopsies of papillary breast
lesions (25 intraductal papillomas and 25
papillary DCIS)

Department of Pathology, Singapore General
Hospital, 2002~2003

Variously reported by 14 general surgical
pathologists - initially

Diagnostic review and assignment to papilloma
and papillary DCIS by Pathologist P.H.T

Results were all in concordance with initial
diagnosis
Materials and Methods
Cont
Intraductal papillomas (25 cases)
Age: mean 44.1 y/o (22-78 y/o)
Epithelial hyperplasia of mild to florid
degree
No atypical ductal hyperplasia

Papillary DCIS (25 cases)
Age: mean 57.4 y/o (35-78 y/o)
No invasive elements present
Cont
43 Hong Kong cases
As to confirm the result with a separate
distinct cohort
Initial diagnosis: 10 general surgical
pathologists
Histologically reviewed by pathologist G.M.T
Cases: age: mean 52.5 y/o
1993 ~ 2001
Excision biopsies 36 cases
Core biopsies 7 cases
Submitted for CK immunohistochemical staining at
SGH, without prior discussion

Materials and Methods
Cont
Immunohistochemistry
Human tonsil, squamous cell carcinoma
and prostate: positive controls for CK5/6,
CK14, and 34BE12, respectively

Normal ducts and ductules in the breast
tissues: internal control
Materials and Methods
Cont
Scoring of sections
Staining intensity: 0, no staining; 1+, weak; 2+,
moderate; 3+, strong

Quantification of positivity (0%~100%)
Estimate of the percentage of stained tumor cells in
the lesion

Immunoscores: multiplying the staining intensity
with percentage positivity (0~300)
Negative or low (0~50); Moderate (51~100); High
(101~200); Very high (201~300)

Materials and Methods
Cont
Confocal microscopy

Statistical analysis
Two-tailed t test: differences in immunoreaction
between the two sample groups
Positive predictive value: CK immunoscore of <50
Papillary DCIS
Negaive predictive value: CK immunoscore of >50
Papilloma

Reevaluate for cases with discrepancies
Materials and Methods
Results:
Cytokeratin expression in a normal breast ductule
Immunoreactions
Intraductal Papillomas and Papillary DCIS (SGH cases)
CK5/CK6
Papillomas
72% : moderate to high immunoscores
Papillary DCIS
All: low immunoscores with 10 being completely negative
CK14
Papillomas
84% high to very high immunoscores
16% moderate to low immunoscores
Papillary DCIS
84% low immunoscores
16% moderate immunoscores
34BE12
Papillomas
56% high to very high immunoscores
44% low to moderate
Papillary DCIS
80% low
20% moderate (1 case: high positivity)
Immunoreactions
Intraductal Papillomas and Papillary DCIS (SGH cases)
t test
Staining intensity, precetage positivity,
immunoscores for each CK:
all three parameters showed significantly
higher in papillomas than DCIS
Immunoreactions
Intraductal Papillomas and Papillary DCIS (SGH cases)
CK5/CK6
CK5/CK6 expression in an intraductal papilloma (left panel) and
Papillary DCIS (right panel)
About half of the tumor cells in the papilloma were stained, whereas
tumor cells in DCIS were nonreactive
CK14
CK14 expression in an intraductal papilloma (left panel) and
Papillary DCIS (right panel)
More than half of the tumor cells in the papilloma were stained,
whereas tumor cells in DCIS were nonreactive
34BE12
34BE12 expression in an intraductal papilloma (left panel) and
Papillary DCIS (right panel)
Approximately half of the papilloma tumor cells were stained,
some DCIS tumor cells were also decorated
Confocal microscopy of CKs
Results
Hong Kong cases
Immunoscores objectively determined by
PHT (SGH)
CK5/CK6 corroborated
Papilloma 89.3%
Papillary DCIS 86.7%
CK14
Papilloma 92.9%
Papillary DCIS 86.7%
34BE12
Papilloma 96.4%
Papillary DCIS 33.3%
Discordant cases
SGH and HK cases
Statistic values
Discussion
Cytoskeleton
Microtubules, microfilaments and
intermediate filaments (CKs belong to one of
5 classes of intermediate filaments)
CKs: cytoplasmic scaffold
Sustain mechanical and nonmechanical stresses
Participation in the response to stress, cell signaling
and apoptosis
To date: 20 CKs (12: acidic type I; 8 neutral-basic
type II)



Discussion
Normal resting mammary gland
Epithelium lining
Inner luminal epithelial (LE) cells
CKs 7, 8, 18, 19
Outer myoepithelial (ME) cells
CKs 5, 14, 17

Discussion
Previous studies
Monoclonal antibodies specific against simple
and/or basal type CKs
Benign and malignant intraepithelial proliferations
of breast
Atypical proliferations
Invasive breast carcinomas

No study compares the expression of basal-
type CKs in papillary tumors

Discussion
Our results
CK14 stained a significantly larger
percentage of tumor cells in papilloma
Breast epithelium of proliferating
mammary gland
3 types of cells: immature precursor (CK5/CK6)
intermediate (CK5/CK6, CK 8, 14, 18, 19), fully
mature (CK14, 18, 19)
Intraductal papilloma
Larger proportion of fully mature cells
Discussion
34BE12
Stained more DCIS tumor cells
Recognizes CK1, 10, 5, 14
CK10 expressed in some LE cells in breast
cancers (previous studies)
Discussion
Results applied to a separate distinct
cohort of HK cases, confirmation of
Benign papilloma diagnosis 89.3% (CK5/CK6)
to 96.4% (34BE12)
Discordant cases: small or core biopsy
ADH case: necessity for microscopic reevaluation

Papillary DCIS (result at odds)
Core biopsy
Invasive component (detect LE as well)
Not as reliable or sensitive as the other 2 CKs
Apocrine nature


In summary
The three CKs can serve as helpful
adjunctive markers
Particularly combination of CK5/CK6 and
CK14
34BE12: low detection rate
Especially in delineation of papillary DCIS

Use of CKs on small lesions, core biopsies,
apocrine morphology, and associated
invasive cancer need further evaluation