HER2 Immunohistochemistry: Workflow Experience with Image Analysis Based Interpretation of

CB11 and 4B5 Clones

Jeffrey Fine MD, Rohit Bhargava MD, Urvashi Surthi PhD, and David Dabbs MD

Magee-Womens Hospital of UPMC

Disclaimer

• One of the abstract authors (David Dabbs) is a consultant for Ventana

• None of the other authors have any conflicts of interest to report

Objectives

• Overview of Her2/NEU testing (IHC)

• Recap of validation results

• Discuss implementation of image analysis

Her2/NEU

• Test for responsiveness to Trastuzumab (Herceptin)

• Trastuzumab is cardiotoxic and is very expensive– False positives are highly undesirable

• Trastuzumab can increase survival or reduce risk of recurrence– False negatives are also undesirable

CAP HER2-A Survey (Spring 2007)

• 40 TMA cores (4 slides)

• Stained/interpreted at institutions (350ish)

• Consensus (>80%) in 22 of 40 cases

• Very variable

IHC Variables

• Pre-analytic– Fixation issues, tissue processing

• Analytic– Validation, Calibration, Antibody clone,

Antigen retrieval, Automation, Controls, etc.

• Post-analytic– Interpretation criteria– QA procedures

• Image analysis (17.9% reported using it)

Validation of IA at UPMC

• Formalin fixed (8-48 hours) paraffin embedded tissue

• Automated IHC platform (Ventana)

• CB11 and 4B5 antibodies (Ventana)

• VIAS (Ventana Image Analysis System)

• FISH (Vysis)

VIAS (image from vendor)

Validation Results I

• System differentiated between tumor and stroma (subjective impression)

• Pathologist had to find invasive tumor (and exclude in-situ tumor)

Classification (Tumor vs. Stroma)

Results CB11

• 100% Concordance with FISH (n=52)– 0/1+ IHC with no amp by FISH– 3+ with amp by FISH

• FISH rate (2+ rate):– Expert 22.9% (n=118)– VIAS 21.2%

• (expert was also 100% concordant)

Results 4B5

• 94.6% Concordance with FISH (n=56)– 100% Concordance with new reference range

• FISH rate (2+):– Expert 21.9% (n=114)– VIAS 28.9% (n=117)

• *new reference range

Reference Range

• VIAS assigns a raw number score to each case which is then rounded to the nearest whole number.

• Out of the box—score 2.5 or higher was rounded to “3+”

• New range is conservative—only cases with score 3.5 are called “3+”

Demixed to show brown

Old Workflow

Order Her2/NEU

Retrieve IHCStain

InterpretStain

Dictate Results

Sign out case

MailboxOffice

New Workflow

Order Her2/NEU

Retrieve IHCStain

Perform IA

Dictate Results

Sign out case

Office MailboxIA Workstation

Implementation Details

• Billing– Fee code 88361– Technical charge initiated by order in APLIS

• Documentation– VIAS results printout retained with other case

paperwork (requisition)– IHC results (ER/PR and Her2/NEU) dictated

into canned text that includes VIAS blurb

Documentation Support

• Transcription– New “quick text” with VIAS sentence– Communication with Transcription Team

• Slide/paper management personnel– Communication – do not discard results report

• Back-up of data– Currently performed manually

“Consumables”

• IA charged per “click”

• IHC Laboratory responsible for keeping an adequate supply of the click reagent– Smart Card (100 tests)

Image from nist.gov www site

Training

• Local Domain Expert (me)

• Fellows

• Selected Faculty

• Other Faculty

• Residents

Status Report

• IA is in production for a week

• Bumps being ironed out– Transcription– Training

• End experience varies

Good

• Should increase standardization– Recent switch to 4B5 clone – different

appearance and possible differences in interpretation

• Documented response to pressure for accurate Her2/NEU testing

• Foot in the door for other IA applications• Vendor has been responsive and appears

to want to improve shortcomings

Bad

• IA of new antibody is less accurate than that of discontinued antibody– Reference range work-around

• Workflow involves “travel” and is more labor intensive than traditional method– Quality improvement but does not extend the pathologist

• Operator error is possible– Data entry– Lighting– Focusing

Future IA (More Automation)

• Whole Slide Images (some systems do permit IA)– Slide could be scanned in the IHC lab, and results

(with the electronic slide) delivered straight to the pathologist

• Automated detection of invasive tumor– Transition of IA to non-pathologist staff

• Electronic interfaces to support test ordering and resulting (no more paper print outs or dictation)

Conclusions

• IA is validated and should improve performance of Her2/NEU IHC testing by reducing post-analytic variability

• Current IA set-up not ideal but an important first step:– Successful implementation in a busy

academic setting– Revenue (digital pathology business case)