Breast Pathology

RCPath Data Sets

Dr Naila Atif
MBBS , FCPS(PK), DipRCPath(UK), FCPP(PK), MIAC
 Reasons for implementing the data Set
• 1-Certain features of Invasive carcinoma are associated with clinical outcome.
• Appropriate treatment
• Screening programmes
• Changing patterns recognition by cancer registries
• 2- Classification of DCIS together with reporting of margins of excision and size
has bee shown to be related with recurrence after local excision and may influence
the use of mastectomy or adjuant radiotherapy
• 3- Radiology / histopathology correlation to ensure , mamaographically detected
lesions have been sampled and accurately diagnosed
• This document also serves to provide guidance for pathologists when
participating in the UK breast pathology EQA scheme.

• The EQA scheme now incorporates a measure of performance appraisal.

• Document has been updated to the standards of the RCPath cancer datasets.
 Key changes in this edition
• Document has been updated to the standards of the RCPath cancer datasets.
• Improved guidance on specimen examination, including handling of oncoplastic and
postneoadjuvant therapy specimens.
• A back-to-basics approach, including recommendations on fixation, macroscopic handling
and measurement of tumour size.
• Recognition of the implications of the 2015 Association of Breast Surgeons’
recommendation of a 1 mm or greater margin distance as definition of complete excision
on pathological specimen handling.
• Guidance on ER/PR/HER2 staining and reporting, including the role of NEQAS, QA,
minimum numbers, audit and benchmarking.
• Clarification of the definition of negative, borderline and positive HER2 status.
• New appendix of routinely used immunohistochemistry adjuncts.
• Guidance and synoptic reporting template for reporting post-neoadjuvant specimens.
• Significant updates to reporting of lymph nodes, adopting a pragmatic approach.
• The term ‘multifocal/multicentric’ replaced by ‘multiple invasive‘.
• Greater emphasis on adherence to criteria use for assessment of tumour type and use of
90% purity rule for definition of pure special type and 50–90% rule for mixed types.
• Further clarification on change of definition of carcinomas with medullary-like features.
 Specimen handling , General Principles
• Type of surgical procedure will be influenced by pre operative diagnosis has been
achieved and the nature of biopsy , Benign, intermediate, (core/cytology, Breast
biopsy B3/C3 or B4/C4) or malignant.
• If no preoperative diagnosis , diagnostic open biopsy with removal of tissue with
minimal surrounding tissue.
• Minimum weight 20gm, should be included in pathology report.
• Impalpable, image guided localization , using wire, dye, radio isotope,
• Frozen section examination is inappropriate for diagnosisof screen detected lesions.
 Benign Preoperative diagnosis.
• If a benign preoperative diagnosis has been made, the lesion may be
removed at the patient’s request. Such resection specimens should be
confined to removal of the lesion with a minimal amount of surrounding
tissue, to avoid leaving a cosmetic defect.
 Mailgnant diagnosis
• If a malignant diagnosis has been made, the surgical procedure is therapeutic.
The type of operation (e.g. wide local excision or mastectomy) will be
influenced by the nature, size and location of the lesion, as well as by patient
choice.
• Before examining the specimen the pathologist should ensure that they are
aware of any previous pathological findings, including the pre-operative
diagnosis, pre-operative clinical and radiological findings, including the
nature, size, site and location of the lesion(s) and any previous treatment (e.g.
neoadjuvant chemotherapy) .
 Surgical Handling
• Resection according to surgical protocol.
• if dissection does not extend to the deep fascia or skin when this is the
norm, this should be clearly indicated on the request form, as this will
influence the examination of the margins of the specimen.
• The surgeon should orientate all breast cancer resection specimens.
• s. Each unit should establish a code of orientation using either different
lengths, or number, of sutures and/or metal staples/clips or ink.

• The nipple extension/direction of the nipple should be separately marked. If

clip/suture placement differs from the agreed local protocol this should be
clearly stated on the request form
• If more than one piece of tissue is removed, it should be made clear (e.g.
using clips and/or diagrams) how the samples are orientated with respect to
each other in order to simplify assessment of the size of the lesion and final
distance to resection margins.
• Intra-operative specimen radiography is mandatory for impalpable lesions
requiring localisation and recommended for all wide local excision
procedures.
• It is strongly recommended that the specimen should be sent immediately to
the pathology laboratory and pre-dissected/incised, ideally in the fresh state.
• . If incision of the fresh specimen is not possible, it should be immediately
placed in an adequate volume of fixative, at least twice that of the specimen.
In the latter circumstance, and by arrangement with the pathologist,
consideration should be given to training the surgeon to make a controlled
single or cruciate pair of incisions into the lesion from the posterior aspect,
thus preserving the integrity of key margins while allowing immediate
penetration of fixative
 Benefits of good fixation
• The benefits of rapid fixation (good tissue morphological conservation with
preservation of mitotic figures and retention of proteins such as oestrogen
receptor) in general outweigh the desire to preserve the specimen intact prior
to examination by the pathologist.
• This is most important for mastectomy specimens into which formalin
penetration can be particularly poor resulting in tumour autolysis with
consequent effects on mitotic count as a component of histological grade,
biomarker expression including oestrogen receptors (ER) and the assessment
of lymphovascular invasion.
 Laboratory handling
• This can be facilitated by prior removal of surface lipid by dipping the
specimen in alcohol and drying and then applying an appropriate pigment
such as India ink, Alcian blue, dyed gelatine or a multiple ink technique.
• Multiple inks can be used.
• India ink can be fixed after painting using 10% acetic acid.
• Specimens must be placed in sufficient formalin (twice the volume of the
specimen)
 Diagnostic localization excision biopsies
• The specimen should be inked, weighed, measured in three dimensions and then, usually, serially sliced at intervals of
approximately 3–5 mm.
• X ray examination , This enables blocks to be taken from the areas corresponding to the mammographic abnormality as
well as any other suspicious areas identified.

• Images can be annotated to indicate sites of block selection.

The sampling technique and the number of blocks taken are clearly dependent on the size of the specimen and the size of
the abnormality. If the specimen is small (e.g. less than 30 mm), it is best to block and examine all of the tissue.
Blocks should be taken to enable a measurement of the histological size of the lesion. Where the maximum macroscopic
dimension of a tumour can be blocked directly, it is recommended that a single block across this aspect be taken.
Where a lesion is larger than can be assessed in a single block, a large block to encompass the maximum dimension may be
taken. When taking large blocks at leastone other normal sized lesional block should be processed as well, to allow optimal
processing and to avoid the excessive use of antibodies in any immunohistochemistry.
• If large blocks are not available, two or more blocks are recommended from
the maximum macroscopic dimension, so that the total tumour size can be
estimated by adding the portions together or measuring the maximum
dimension on the two slides fitted together
• For diffuse tumours, especially diffuse lobular carcinomas, it may not be
possible to define macroscopically the true extent of tumour and in this case,
either a large block or consecutive blocks of the whole abnormal area
(including adjacent fibrotic tissue) may be necessary.
• For larger specimens, sampling should be adequate to determine accurately
the size of the lesion.
• If specimens are sent as more than one piece of tissue, it can be impossible
to measure the absolute extent of the lesion. In these cases, it is appropriate
to take a pragmatic approach and to measure the maximum size in each piece
of tissue and add the dimensions to give an estimated total size.
• Occasional cases will have had a diagnostic excision biopsy before definitive
treatment, or primary chemotherapy or exceptionally a frozen section may
have been performed.
• Tumour size assessment in these circumstances may be necessarily inaccurate
and an evaluation based on the ultrasound or radiographic size in
conjunction with the histology may be necessary.
• Although pathology measurement of tumour size is considered the ‘gold
standard’,
 Therapeutic wide local excisions
• Lesions that have a pre-operative diagnosis of malignancy and are deemed to be
suitable for breast conserving surgery with regard to clinical/radiological size may
be excised as a therapeutic wide local excision.
• The specimen should be weighed and measured in three dimensions.
• The specimen should have been incised to allow prompt fixation (see above and
Figure 2a) and excision margins should be inked and the specimen can be sliced
either before fixation or (less preferably) after fixation.
• . Whichever is utilised, as an absolute minimum, the information for the breast
cancer dataset, including accurate measurement of size and detailed examination of
the margin status and distance to margins, must be provided.
 For all methods
• Details of the macroscopic appearances of the specimen should be recorded
including: tumour size in three dimensions distances to all margins.

• The number of tumour blocks will vary with tumour size but is usually at
least three. The edges of the tumour with surrounding uninvolved tissue
should also be examined in all three dimensions to identify associated DCIS
and peritumoural lymphovascular invasion not visible to the naked eye, and
permit accurate an assessment of whole tumour size.
• If therapeutic samples are sent in more than one portion, it can be extremely
difficult to measure the absolute largest extent of the whole lesion present.
In these cases it is appropriate to measure the maximum distance in any piece
of tissue and to add the dimensions to give an estimated total size or
preferably defer to the imaging size. If, however, the orientation of the
specimens can be determined, the size can be ascertained more reliably
• All surgically relevant margins of therapeutic excision specimens should be
sampled. This will include all radial/circumferential margins (superior,
inferior, medial, lateral and nipple margins), and the deep (posterior) and
superficial (anterior) margins if dictated by local protocol. Particular
attention should be paid to the margin nearest the abnormality and the
margin nearest the nipple.
• The use of different colour inks/markers on an individual section can assist
microscopic identification of specific margins.
1.6.2 Wide local excisions for ductal carcinoma
in situ (DCIS): presenting as mammographic
calcification
• DCIS typically presents as a mammographically detected abnormality, usually calcification,
which may not be visible on macroscopic examination of the sliced tissue.
• It is usual for the surgeon when performing a therapeutic operation to take all of the tissue
from the subcutaneous aspect to the pectoral fascia.1 It is essential that the pathologist is
informed if the usual surgical protocol has not been undertaken as this will affect the
optimum specimen handling methodology, e.g. central excisions, or specimens where breast
tissue remains at the deep (posterior) and superficial (anterior) aspects of the excision, and
the distance to these margins is thus clinically relevant.

• , the surgeon should mark the nipple duct margin; DCIS tracks towards the nipple4 and, in
this plane in particular, can be some distance from the obvious area of microcalcification.
• The specimen excision margins should be inked and the specimen can be
sliced either before or after fixation. The use of different colour
inks/markers on an individual section can assist microscopic identification
of specific margins. Inks which are radio-opaque should ideally be avoided if
applied prior to slice X-ray. If the specimen is large, then incision before
fixation is recommended. The specimen should be sliced at intervals of
approximately 3–5 mm
• Serial slicing enables specimen slice radiographic mapping of the specimen
which provides a high level of confidence that the lesion has been accurately
and adequately sampled; slicing and X-raying the specimen slices enables
blocks to be taken most accurately from the areas corresponding to the
mammographic abnormality as well as from any other suspicious areas
identified. This is essential to avoid underestimation of lesion size and
overestimation of the distance to specimen margins.
• Macrophotography or schematic diagrams may also assist in recording macroscopic
findings and the block map as well as identifying individual sampled margins.

• Sampling may be facilitated by the identification of any radiological marker (e.g. clip,
collagen marker or coil). Tissue changes relating to previous core biopsy are an
important landmark to indicate sampling of the site of the index lesion and should
be recorded in the report, particularly if the whole abnormality was removed by the
cores. The macroscopic and or radiographic lesion should be described and its
size in three dimensions and distance to margins recorded.
 Number of blocks

• The number of blocks taken will depend on the size of the

specimen and the size of the abnormality. If the specimen is
small, or if slice radiology unavailable, it is best to block and
examine all of the tissue.
• For larger specimens sampling should include the extremes of the
radiographic calcification and adjacent tissue in order to avoid
underestimation of the size of the lesion.
• it is recommended that one to two block per 10 mm of the maximum
dimension of the area of calcification be taken.

• Measurement can be made in this way from the most distal involved duct
across the main area of calcification to the most proximal involved duct
 Margins
• All surgically relevant margins of therapeutic excision specimens should be
sampled. This will include all radial/circumferential margins (superior,
inferior, medial, lateral and nipple margins), and the deep (posterior) and
superficial (anterior) if dictated by local protocol or by the surgical procedure
from an individual patient.
• Particular attention should be paid to the margin nearest the mammographic
abnormality and the margin nearest the nipple.
 Margin in multiple lesions
• If therapeutic samples are sent in more than one portion, it can be extremely
difficult to measure the absolute largest extent of the whole lesion present.
In these cases, it is appropriate to measure the maximum distance in any
piece of tissue and to add the dimensions to give an estimated total size. If,
however, the orientation of the specimens can be determined, the size can be
ascertained more reliably.
 1.6.3 Cavity shave/biopsy specimens
• Surgical resection margins. This is typically done after taking the radial
tumour blocks. This can produce a series of additional blocks including:
superior shave, supero-lateral shave, lateral shave, infero-lateral shave, inferior
shave, infero-medial shave, medial shave and supero-medial shaved edge,
depending on the size of the specimen.
• it is recommended that shave margin specimens <1mm thick are sliced
perpendicular to the new margin face so that the distance to margins can be
recorded.
 Bed biopsies
• The surgeon may provide separate cavity shaves, which may be submitted to
the laboratory as ‘bed biopsies’. The site of each specimen should be clearly
labelled and each specimen examined separately.
 Re-excision specimens
• Re-excision specimens can be weighed and serially sliced at 3–5 mm.
• Blocks taken should be recorded in such a way as to permit accurate
assessment of the adequacy of excision and size of any malignant lesions
identified.
• It is difficult to be proscriptive regarding the extent of block sampling as
the nature and size of these specimens varies; the focus should be on the
new excision margin rather than exhaustive detection of residual disease.
• If re-excision specimens have been taken which contain further tumour, it
can be extremely difficult to determine the absolute size of lesion. A
pragmatic approach is required, and the maximum distance in each piece of
tissue can be measured and added to give an approximate total size of
tumour. If, however, the orientation of the specimens can be determined, the
size of tumour can be ascertained more reliably.
 Mastectomy specimens
• 1.7.1 Mastectomy specimens for invasive carcinoma: presenting as mass lesion (Figures 5a and
5b)
• The tumour is conventionally incised from the deep (posterior) fascial plane in the
• sagittal plane at a maximum of 10 mm intervals after inking, e.g. with India ink (Figure
• 5). Differential colour marking of anterior, posterior and radial surfaces may facilitate
• orientation both prior and subsequent to block taking in skin-sparing mastectomies.
• Slicing in the coronal plane from deep (posterior) to anterior (superficial) (Figure 6) may
• be appropriate in some cases, particularly where it may facilitate correlation with
• imaging findings.
Centre of the tumour may be incided in cruciate fashion

• Normal breast should be sliced at 10n mm interval

•
Tumour size in three dimensions
• Multiple tumours , distance between tumours should be sampled.
• Tumour blocks – multiple
• Edges of tumour with uninvolved tissue
• Numbur of blocks may vary
• Large blocks are helpful
• Lateral edge of the specimen should be palpated for intramammary lymph nodes, low axillary lymph nodes
• Nipple examination
• Sections from nipple areola complex.
• Nipple --- coronal section
• Occult paget disease === sagittal section
•
 Mastectomy specimen for DCIS ,
mammographic calcification
• Combined radio, histo correlation is must
• All mastectomy specimen should be oriented , by placing a suture in axillary tail
• Posteror aspect is painted
• Specimen is incised at 10mm interval
• Can remove nipple separately before fixation
• X ray should be done after fixation
• Accurate localization of lesion
• Minimum of 1-2 blocks per 10mm calcification
• Sampled thoroughly to exclude invasive focus
• Large blocks
• Normal portion 10 mm slice and examine by eye
• Lateral end for intramammary lymph nodes
• Nipple areola complex
• Additional sampling of quadrants
 Completion mastectomy after WLE
• Fixed
• Inked
• Cavity --- granulation tissue, fibrosis, fat necrosis
• Sections wall of cavity
• Nipple
• Other quadrants
 Mastectomy after neoadjuant therapy
• Tumour downsizing
• Difficult tumour localization of there is complete response
• Previous information is very important
• Multiple invasive foci can be missed
• Initial Handling is same
• Adequate and prompt fixation
• Tumour size before treatment important for MDT
• Palpate and take slices , if size is less, take more blocks from periphery
• Significant tumour response --- pale , ill defined, soft , edematous area of
fibrosis
• Residual microcalcification can help in location of tumour
• Whole tumour response should be sampled
• Margin should be sampled thoroughly
• Lymph nodes should be sampled , however the yield can be less.
Oncoplastic specimens
• Contralateral breast reduction specimens from patients with breast cancer
and prophylalactic mastectomies should be sampled more thproughly.
• Manual palpation with 5-10 mm interval slicing
• Minimum two tissue blocks
 Lymph node examination
• Axillary lymph node clearance
• Axillary lymph node samples
• Sentinal lymph node
 Sentinal lymph node
•
examine each lymph node
• Don’t damage the capsule
• Representative complete section of lymph node is advised
• Lymph nodes greater than 4mm should be sliced at interval of 2mm perpendicular to long axis
• Lymph nodes less than 4mm sliced and submitted entirely
• Examination of levels is not necessary
• Immunohistochemistry AE1/AE3, broad spectrum cytokeratins
• Dendritic cells and lymphoid cells can give fasle positive results

•
 Axillary clearance specimens
• All lymph nodes should be sampled and involved and uninvolved lymph
nodes should be stated in report.
• Bouin solution, clearing solution may increase the yield
• Each lymph node should be identified and examined
 Intraoperative examination of lymph nodes

• Not as a routine
 Frozen section and Molecular Techniques
• Detects 70% of metastasis
• Touch imprints 63 %
• One step nucleic acid amplification OSNA, approved by nice , 2013
• RD100i OSNA system
 6- CORE DATA ITEMS
• Side
• Pathologist
• Date
• Radiograph
• Mammographic abnormality
• Histological calcification
• SPECIMEN TYPE
• WLE
• Excision
• Diagnostic localization
• Segmental excision
• Mastectomy
• Re- excision
• Futrher margins
• Duct Excision specimens
• Sentinal lymph node
• Axillary sampling
• Axillary lymph node excision
• Vacum assissited excision
• Specimen weight
• Benign / malignant
 Tumour classification and prognostic factors

• Tumour size
• Invasive tumour size
• Satellite tumour nodules should not be measured in main tumour dimension
• Features that may be assistance to separate two deposits whether , satellite or
main tumour , a distance of 5mm or more will separate as a second focus
 Whole tumour size,invasive tumour and
surrounding DCIS
• The measurement of DCIS with invasive carcinoma should be recorded in
the whole tumour size fieldon reporting form, including tumours which are
composed of predominantly DCISbut have multiple foci of invasion.
• Invasive tumour size should be given in invasive component.

• Pathologist should take blocks between tumour and the excision margin
 Insitu carcinoma
• DCIS
• Pleomorphic lobular carcinoma
• Classical lobular ---- multifocal , measurement is un necessary.
• Pleomorphic lobular--- focal , can give to ipsilateral invasive carcinoma.
 Disease Extent
• The designation of multiple foci should be reserved for multiple separate
areas of invasive tumour, such as occurs with invasive lobular carcinoma or
tumours with extensive lymphovascular invasion where there are multiple
areas of invasive tumour due to extravasation of tumour cells from
lymphatics and establishment of separate satellite invasive tumour foci.
 Histological Grade
• Tubule/acinar formation Score 1. >75% of tumour forming tubular structures 2. 10–75% of
tumour 3. <10% of tumour.
• 6.2.2.2 Nuclear atypia/pleomorphismScore 1. Nuclei small with little increase in size in
comparison with normal breast epithelial cells, regular outlines, uniform nuclear chromatin, little
variation in size 2. Cells larger than normal with open vesicular nuclei, visible nucleoli and
moderate variability in both size and shape. (Figures 20e, 32d) 3. Vesicular nuclei, often with
prominent nucleoli, exhibiting marked variation in size and shape, occasionally with very large
and bizarre forms.
• 6.2.2.3 Mitoses Score
•
• The mitosis score depends on the number of mitoses per 10 high power fields
• 6.2.2.4 Overall grade
•
• The use of terms such as well differentiated or poorly differentiated in the
absence of a numerical grade is inappropriate. The scores for tubule
formation, nuclear pleomorphism and mitoses are then added together and
assigned to grades, as below: Grade 1 = Scores of 3–5 Grade 2 = Scores of
6 or 7 Grade 3 = Scores of 8 or 9.
 6.2.4 Lymph node stage

• Stage 1: Node negative; Stage 2: 1–3 nodes positive; Stage 3: 4 or more

nodes positive.
 Micrometastasis , isolated tumour cells
• Micrometastasis is defined as one or more deposits of metastatic carcinoma within
the lymph node or the node capsule that are more than 0.2 mm in size but none of
which is larger than 2 mm in greatest dimension. Lymph nodes involved by
micrometastases are regarded as positive.
• Cases with only isolated tumour cell clusters (ITCs) in regional lymph nodes are
classified as node negative (pN0). ITCs are single tumour cells or small clusters of
cells not more than 0.2 mm in greatest dimension (Figure 43c) or single tumour
cells, or clusters of fewer than 200 cells in a single histological cross section. These
may be detected by routine H&E, by immunohistochemistry or molecular methods
but which may be verified on H&E stains. ITCs do not typically show evidence of
metastatic activity (e.g. proliferation or stromal reaction).
• It is therefore recommended that any lymph node involvement >0.2 mm but
<2 mm in any of the three dimensions is categorised as a micrometastasis.
The 0.2 mm size cutoff relates to the maximum diameter of the largest
tumour cell cluster. There may be instances of nodal involvement with the
largest cluster measuring <0.2 mm in diameter but containing >200 cells, and
vice versa, i.e. clusters >0.2 mm in diameter with <200 cells. Size should be
considered first, and the cell count applied only if the largest cluster is <0.2
mm. 66
• any residual tumour cells identified in a lymph node examined after
neoadjuvant treatment should be considered as positive
 Excision margin
• > 5mm is considered safe
 Recognising carcinoma after neoadjuvant
therapy
• Carcinoma cells retain cytokeratin expression post-therapy, whilst
macrophages will continue to express CD68. There is some evidence that
AE1/AE3 may be preferable to other cytokeratin markers in the
immunohistochemical examination of sentinel lymph nodes out with the
setting of primary systemic therapy6 but reticulum and inflammatory cells
(which may give positive reactivity with Cam5.2 and other pan-cytokeratin
formulations) may be especially difficult to assess in a background of
neoadjuvant chemotherapy changes.
• Table 7: The Allred/Quick score and H score methods for hormone receptor IHC
semi-quantitative scoring
•
• Allred score (0–8 Quick score) 81 Score for proportion Score for intensity 0 = no
staining 0 = no staining 1 = < 1% nuclei staining 1 = weak staining 2 = 1–10%
nuclei staining 2 = moderate staining 3 = 11–33% nuclei staining 3 = strong
staining 4 = 34–66% nuclei staining 5 = 67–100% nuclei staining The scores are
summed to give a maximum of 8. The cut off for positivity using Allred score ≥3
 Quality assurance for oestrogen receptor
evaluation
• Participating laboratories are requested to stain the UK NEQAS ICC and ISH
section using their everyday clinical methodology. As well as the UK NEQAS ICC
and ISH slide, participants are also requested to submit their own in-house control
material for assessment. It is strongly encouraged that in-house control material
should comprise three breast carcinomas showing the following level of expression
(i) >80% tumour positivity with high intensity (Allred/Quick score 7–8) (ii) 30–70%
tumour positivity with low-moderate intensity (Allred/Quick score 4–6) and (iii) a
negative tumour, with positively stained normal glands (Allred/Quick score 0).
Participants are also requested to complete a web-based methodology form,
including such information as antibody clone and dilution, heat mediated antigen
retrieval method, automation staining instrument, etc.
 Assessment procedure

• Participants then return their stained slides to the organising centre, for
evaluation by a panel of four expert assessors. Each of the four assessors
awards marks out of 5, which are then totalled to give a score out of 20
Assessment of human epidermal growth factor
receptor 2 (HER2)
• Thank u