Should we still be performing IHC on all sentinel nodes?

Miami Breast Cancer Conference 31 st Annual Conference March 8, 2014 Should we still be performing IHC on all sentinel nodes? Donald L. Weaver, MD Professor of Pathology University of Vermont USA

Objectives Participants should be able to: Define occult metastases and understand their value for answering research questions Understand that we cannot practically identify all metastatic tumor present in SLNs Understand that IHC identifies the least significant (smallest) metastases Identify clinically useful strategies for detecting LN metastases based on statistical principles No financial disclosures to declare

A more fundamental question Q: Should we have ever started using IHC for sentinel node evaluation? A: Yes and No Yes No IHC has been a powerful research tool IHC is an arbiter of questionable findings IHC was not scientifically validated for prognosis or prediction [Do not use on all cases in general practice]

Why did we use IHC?

Proof of SLN hypothesis in a blinded analysis of occult metastases detected in SLNs and nonslns: Metastases are 12.3 times more likely in SLNs than nonslns (p<0.001) Cancer 2000;88:1099-1107 Additional nonsln metastases 13.4 times more likely when SLN is positive

Defining Occult Metastases An occult metastasis is any metastasis that is either missed or not identified on initial examination using a pre-defined or standard evaluation protocol Occult metastases facilitated prospective blinded analysis of the prognostic significance of isolated tumor cell clusters (ITCs) and micrometastases

Studies of Occult Metastases All studies of occult metastases whether retrospective or prospective are asking the same fundamental question: How extensively should pathologists evaluate lymph nodes for metastatic disease? Guiding principle: Outcome NOT prevalence Survival Disease free survival Axillary recurrence

IHC can detect single tumor cells THEREFORE to assign importance to single cell detection, single cells would have to be systematically excluded from all negative nodes

Lymph Node Evaluation Strategies Objective: Identify all single tumor cells Typical single tumor cells in SLNs measure 10 microns (0.01 mm) Technical cost assumptions: 2 SLNs per patient, 1 block per SLN Slice SLNs at 2.0 mm, embed all slices 200 CK IHC slides per block at US $12 each (cost) 144,000 new node negative breast cancers annually

Detecting single tumor cells in SLNs: Can we afford it? No! 57.6 million slides to screen (76 pathologist yrs @ 10 sec/slide) US $691 million annually (Does not include interpretative charges)

Cytokeratin Immunohistochemistry DOES NOT guarantee the pathologist will not miss isolated tumor cells and clusters

0.01 0.02 0.03 0.04 0.05 0.06 0.07 0.08 0.09 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 number of cases B-32 Occult SLN metastases identified by pathologist (LM) and computer assisted cell detection with image analysis (CACD) 10 9 8 7 6 5 4 3 2 1 0 MISSED 75% ITCs missed CACD only LM 47% Micromets missed 236 cases screened 34/236 (14.4%) pos by LM 30/202 (14.9%) pos by CACD Prevalence of additional mets Pathologists will miss isolated CK positive tumor cell clusters that are no larger than 100 microns (0.1 mm) largest occult metastasis (mm) Cancer 2006; 107:661-667

Do not be impressed by what pathology has found Understand what the evaluation has missed

NSABP B-32 trial investigation of micrometastases A B Clinical protocol used for patient management Nodes sliced at 2 mm intervals One section per block IHC only for suspicious H&E findings Blinded occult metastasis study of negative SLNs Two additional levels: 0.5mm and 1.0mm deeper Two sections at each level: H&E plus CK IHC C Quality assurance pilot study Additional levels every 0.18 mm through block CK IHC at every level Results compared to two-levels at 0.5 mm and 1.0 mm

NSABP B-32 node positive rates slice all nodes at 2mm prior to embedding; embed all sections (calculations on per case basis) spacing 0.5mm 0.18mm macromet (%) - 0.4 0.4 micromet (%) - 4.1 6.1 ITC (%) - 10.2 17.4 occult rate (%) reference 14.7 23.9 overall positive (%) 26 40.7 49.9 effort index 1x 3x 12x Data adapted from: Am J Surg Pathol 2009; 33:1583-1589.

The more you look the more you find BUT diminishing returns more slice work all nodes at 2mm prior to embedding; embed all sections (calculations on per case basis) what value to the woman? NSABP B-32 node positive rates spacing 0.5mm 0.18mm macromet (%) - 0.4 0.4 micromet (%) - 4.1 6.1 ITC (%) - 10.2 17.4 occult rate (%) reference 14.7 23.9 overall positive (%) 26 40.7 49.9 effort index 1x 3x 12x $$$ Data adapted from: Am J Surg Pathol 2009; 33:1583-1589.

NSABP B-32 trial investigation of micrometastases A B Clinical protocol used for patient management Nodes sliced at 2 mm intervals One section per block IHC only for suspicious H&E findings Blinded occult metastasis study of negative SLNs Two additional levels: 0.5mm and 1.0mm deeper Two sections at each level: H&E plus CK IHC C versus Quality assurance pilot study Additional levels every 0.18 mm through block CK IHC at every level Results compared to two-levels at 0.5 mm and 1.0 mm

9C 9D 10B 10C 16B1 24B 25B 29A 38B1 38B2 47B1 48B1 49A1 49A2 49A3 50B 52C 52D2 53A2 54A1 metastasis size (mm) Comparison of maximum metastasis size two-level wide spaced (0.5mm spacing, B-32) versus multi-level narrow spaced through block (0.18mm spacing) 0.7 0.6 B-32 comprehensive Two-level protocol (fixed 2 sections) 0.5 0.4 0.3 0.2 0.1 0 2.2% risk of missing micrometastases 8.9% risk of missing isolated tumor cell clusters (ITC) 22% risk that detected ITC is actually a micrometastasis case and block label Am J Surg Pathol 2009; 33:1583-1589 Multi-level protocol (median 11 sections)

N Engl J Med 2011; 364:412-421

NSABP B-32 trial investigation of micrometastases A B C Macromets virtually excluded by design Clinical protocol used for patient management Nodes sliced at 2 mm intervals One section per block IHC only for suspicious H&E findings Blinded occult metastasis study of negative SLNs Two additional levels: 0.5mm and 1.0mm deeper Two sections at each level: H&E plus CK IHC NO TREATMENT BIAS Quality assurance pilot study Additional levels every 0.18 mm through block CK IHC at every level Results compared to two-levels at 0.5 mm and 1.0 mm

Distribution of occult metastases, B-32 3887 cases with initially negative SLNs n 14 172 Macrometastases identified in less than 0.4% of total cases on deeper levels 430 0 100 200 300 400 500 ITC micromet macromet Sectioning SLNs at 2.0 mm intervals effectively excludes virtually all macrometastases from node negative group

Subset analysis of outcome by size of detected occult metastases Occult metastasis size Hazard Ratios Death Second event Distant disease Breast cancer death None detected reference reference reference 1.6% ITC 1.27 1.18 1.19 2.2% Micro/macro 1.60 1.38 1.41 4.0% Nodal tumor burden is a continuous variable size of metastases, number of metastases, and number of involved nodes all contribute unfavorably to outcome total volume of metastatic disease is far more important than simple presence or absence of disease Stratification of nodal tumor burden is justified (AJCC/UICC N-classification)

Probability of Survival Kaplan-Meier Survival Estimates according to presence or absence of occult metastases Occult metastases are a STATISTICALLY significant independent prognostic variable (large studies can detect small differences in outcome with high confidence) The magnitude of the differences in outcomes do not justify changes in clinical management Overall survival Disease free survival Distant disease free 1.2% (p=0.03) 2.8% (p=0.02) 2.8% (p=0.04) OS 5-year K-M estimates OM not detected: 95.8% OM detected: 94.6% DFS 5-year K-M estimates OM not detected: 89.2% OM detected: 86.4% DDFI 5-year K-M estimates OM not detected: 92.5% OM detected: 89.7% n = - + 3197 598 3085 581 2295 416 575 108 n = - + 3092 574 2897 539 2115 375 520 92 n = - + 3122 584 2942 553 2159 383 537 95 Time (months) Time (months) Time (months) NEJM 2011; 364:412-421

Effect of occult metastases on survival Kaplan-Meier 5-year estimates of survival (%) Occult metastases Detected Not detected Delta p-value Overall survival (%) B-32 95.8 94.6 1.2 0.03 Z0010 95.7 95.1 0.6 0.64 Disease free survival (%) B-32 89.2 86.4 2.8 0.02 Z0010 92.2 90.4 1.8 0.82 Median follow-up 7.9 years (B-32) and 6.3 years (Z0010) Prevalence 15.9% (B-32) and 10.5% (Z0010)

Prevalence of occult metastases B-32 Z0010 5611 Randomly assigned 5119 19.2% Tumor >2 cm 12.7% 75.6% Age >50 years 71.1% 7.9 yrs Median follow-up 6.3 yrs 3889 (71.1%) SLN negative 3904 (76.3%) 3884 Blinded evaluation 3326 616 (15.9%) Occult metastases 349 (10.5%) Prevalence directly related to detection strategy NEJM 2011; 364:412-421 JAMA 2011; 306:385-393

Occult metastases (OM) are NOT discriminatory predictors of outcome! Regional and Distant Recurrences 138 (3.6%) of 3884 30 (21.7%) with OM 108 (78.3%) without OM Disease Free Survival 616 of 3884 with OM 496 (80.5%) disease free 120 (19.5%) second event* If additional treatment was given based exclusively on the presence of occult metastases 80% of these patients would be over treated Occult metastases were observed in unfavorable (>2cm) and favorable (ER+) breast cancers * includes any death, contralateral breast cancer, regional and distant recurrence, and second non-breast cancer

Health care dollars for SLN IHC 230,000 new cases of breast cancer per year 172,500 clinically node negative cases (75%) 448,500 sentinel nodes (average 2.6 per case) $100 insurance reimbursement (actual collected) $44.8 million per year

% Surviving NSABP B-32 Sentinel Node Negative Patients Overall Survival by Occult Metastases Status 100 80 Entire Cohort, OS 60 40 20 No Occult Mets 3268 pts., 378 deaths ITC 431 pts., 61 deaths, HR=1.22 Micromets 171 pts., 28 deaths, HR=1.43 0 Data as of Dec 31, 2012 0 2 4 6 8 10 Years after Randomization

% Disease-Free NSABP B-32 Sentinel Node Negative Patients Disease Free Survival by Occult Metastases Status 100 80 Entire Cohort, DFS 60 40 20 0 No Occult Mets 3268 pts., 738 events ITC 431 pts., 122 events, HR=1.29 Micromets 171 pts., 46 events, HR=1.23 Data as of Dec 31, 2012 0 2 4 6 8 10 Years after Randomization

NSABP B-32 (SLN vs AD) Overall Survival with Occult Metastases 86.7% Cohort stratified by treatment arm, OS 85.2% Trt N Deaths SNR+AD 316 47 SNR 300 43 HR=0.98 p=0.91 Data as of Dec 31, 2012

NSABP B-32 (SLN vs AD) Disease Free Survival with Occult Metastases 76.4% Cohort stratified by treatment arm, DFS 69.9% Trt N Events SNR+AD 316 94 SNR 300 76 HR=0.82 p=0.2 Data as of Dec 31, 2012

B-32: SLN-Negative Patients with Occult Metastases Cumulative Incidence Local-Regional Recurrences SNR+AD SNR 316 pts., 18 Local-Regional events 300 pts., 14 Local-Regional events 6.3% HR=0.8 p=0.52 4.2% Data as of Dec 31, 2012

Conclusions Occult metastases are a STATISTICALLY significant independent prognostic variable Micrometastases are more significant than ITCs Overall effect on outcome is small (1-3%) SLNB only arm (highly relevant): 0.5% difference in event free rate 1.3% difference in combined regional and distant recurrence Data do not support use of deeper levels and IHC Occult metastases are NOT discriminatory predictors of any outcome measure 10-year outcome analysis planned

Final Recommendation Thin gross sections (2 mm) Embed and examine each slice 2.0 mm Examine one section from each block Levels not required IHC not required [use levels and IHC sparingly with discretion]