GENOMIC TESTS FOR BREAST CANCER: FACT, MYTH, AND EVERYTHING IN BETWEEN Adam Brufsky, MD, PhD Professor of Medicine Associate Chief, Hematology-Oncology Associate Director, Clinical Investigation University of Pittsburgh Cancer Institute University of Pittsburgh
DISCLOSURES Consulting fees from Agendia Genomic Health Biotheranostics Nanostring Technologies
Why are we here? Oncologists have a lot of powerful tools Many of them are very toxic Some of them are of great benefit Toxicity and benefit often do not correlate We try to understand natural history of cancer We try to understand benefit of intervention Goal: Do what is right for the patient
Clinical Prognosticators-Adjuvant! online
Clinical Predictors: PREDICT-PLUS http://www.predict.nhs.uk PREDICT Plus: development and validation of a prognostic model for early breast cancer that includes HER2. Wishart GC, et al. Br. J. Cancer 2012;107(5):800-7.
Tumor Grade Rakha et al. Breast Cancer Res 2010, 12: 207.
Reproducibility of tumor histological grade Kappa: 0.43-0.83 for inter-observer variability Despite the objective improvements that have been made to breast cancer grading methods, any assessment of morphological characteristics inevitably retains a subjective element and is heavily dependent on the pre-analytical parameters. Rakha et al. Breast Cancer Res 2010, 12: 207.
How Good are Clinical Predictors They are not useless However, there is some subjectivity in certain important measurements, such as grade What else can be done to reduce subjectivity?
Oncotype DX (RT-PCR Technology) 16 Cancer and 5 Reference Genes PROLIFERATION Ki-67 STK15 Survivin Cyclin B1 MYBL2 INVASION Stromolysin 3 Cathepsin L2 HER2 GRB7 HER2 CD68 ESTROGEN ER PR Bcl2 SCUBE2 GSTM1 BAG1 REFERENCE Beta-actin GAPDH GUS RPLPO TFRC RS Weighting: + 0.47 x HER2 Group - 0.34 x ER Group +1.04 x Proliferation Group + 0.10 x Invasion Group + 0.05 x CD68-0.08 x GSTM1-0.07 x BAG1 Category RS (0 100) Low risk RS < 18 Intermediate risk RS 18 and < 31 High risk RS 31
Recurrence Score and Distant Recurrence-Free Survival Rate of Distant Recurrence at 10 years 40 35 30 25 20 15 10 5 Low RS < 18 Rec. Rate = 6.8% C.I. = 4.0% - 9.6% Intermediate RS 18-31 Rec. Rate = 14.3% C.I. = 8.3% - 20.3% High RS 31 Rec. Rate = 30.5% C.I. = 23.6% - 37.4% Recurrence Rate 95% C.I. 0 0 5 10 15 20 25 30 35 40 45 50 Recurrence Score Paik.S. et al. N Engl J Med 2004;351:2817-26
High RS Result Correlates with Greater Benefit from Chemotherapy (NSABP B-20) Proportion without distant recurrence 1.0 0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 All patients RS < 18 RS 18-30 RS 31 Tamoxifen + chemotherapy Tamoxifen Tamoxifen + chemotherapy Tamoxifen Tamoxifen + chemotherapy Tamoxifen Tamoxifen + chemotherapy Tamoxifen N 424 227 218 135 89 45 117 47 Events 33 31 8 4 9 4 13 18 P = 0.02 P = 0.61 P = 0.39 P < 0.001 PATIENTS WITH HIGH RS 28% absolute benefit from tamoxifen + chemotherapy 4.4% absolute benefit from tamoxifen + chemotherapy 0 2 4 6 8 10 12 Years RS, Recurrence Score result Paik S, et al. J Clin Oncol. 2006;24:3726-3734. 11
SWOG 8814: Breast Cancer-Specific Survival of Node-Positive Patients by Treatment and RS Group BREAST CANCER-SPECIFIC SURVIVAL BY TREATMENT RS < 18 RS 18-30 RS 31 100 100 100 75 75 75 50 25 Stratified log-rank P = 0.56 at 10 years CAF T (n = 91, 10 events) Tamoxifen (n = 55, 4 events) 0 0 2 4 6 8 10 Years since registration 10-yr BCSS T: 92% vs CAF T: 87% No benefit to CAF over time for low Recurrence Score 50 25 Stratified log-rank P = 0.89 at 10 years CAF T (n = 46, 10 events) Tamoxifen (n = 57, 11 events) 0 0 2 4 6 8 10 Years since registration 10-yr BCSS T: 70% vs CAF T: 81% Interaction P = 0.021 50 25 Stratified log-rank P = 0.033 at 10 years CAF T (n = 47, 18 events) Tamoxifen (n = 71, 20 events) 0 0 2 4 6 8 10 Years since registration 10-yr BCSS T: 54% vs CAF T: 73% Strong benefit to CAF over time for high Recurrence Score RS, Recurrence Score result 12 Albain KS, et al. Lancet Oncol. 2010;11(1):55-65.
The 21 gene assay: Savior, or a Devil in the Details? Data is very compelling risk of recurrence, for both N(-) and post-meno N(+) Most important data for the practicing oncologist is in chemotherapy benefit prediction the data is also compelling, but low N N = 164 in B20 N(-) high RS group tam +/- chemo N = 118 in E8814 N(+) high RS group tam+/- chemo What exactly is the RS measuring?
Are we simply measuring mrna of genes that we can easily detect by good IHC? 16 Cancer and 5 Reference Genes PROLIFERATION Ki-67 STK15 Survivin Cyclin B1 MYBL2 INVASION Stromolysin 3 Cathepsin L2 HER2 GRB7 HER2 CD68 ESTROGEN ER PR Bcl2 SCUBE2 GSTM1 BAG1 REFERENCE Beta-actin GAPDH GUS RPLPO TFRC RS Weighting: + 0.47 x HER2 Group - 0.34 x ER Group +1.04 x Proliferation Group + 0.10 x Invasion Group + 0.05 x CD68-0.08 x GSTM1-0.07 x BAG1 Category RS (0 100) Low risk RS < 18 Intermediate risk RS 18 and < 31 High risk RS 31 If you are really, really good at IHC in breast cancer, can you replicate the RS info?
The Magee Equations Examined a test set of 817 ER (+), N (-) cases from MWH from 2004-2009 with RS results Derived three regression equations based on ER, PR, Her2, Ki67 scores as well as clinical variables on these cases Blinded test on another set of 255 ER (+), N(-) cases from MWH with known RS results Klein et al Modern Pathol (2013) 1-7.
The Magee Equations Klein et al Modern Pathol (2013) 1-7.
The Magee Equations R=0.61 Klein et al Modern Pathol (2013) 1-7.
Using the Biology of Tumor Pathways: Mammaprint and PAM50
MammaPrint: A 70 Gene Breast Cancer Prognosis Profile 70 significant prognosis genes Tumor samples van t Veer et al., Nature 415, p. 530-536, 2002
MammaPrint interrogates multiple genomic pathways (no ER/PR/Her2 mrna) 1. 2. IGFBP5, TGFB3, FGF18, ESM1, RARRES3, PITRM1, EXT1, EXTL3, SCUBE2, EBF4,CDC42BPA, CDCA7, CDCA7L, GMPS, MELK, RFC4, WISP1, HRASLS, BBC3, DTL, FBXO31, EGLN1, GNAZ, MTDH, FLT1, ECT2, DIAPH3, NUSAP1, AKAP2, NDC80, PRC1, ORC6L, CENPA, DCK, CCNE2, MCM6, QSOX2, STK32B COL4A2, FLT1, FGF18, MMP9 3. FLT1, TGFB3, IGFBP5, FGF18, RARRES3, CDCA7L, WISP1, DIAPH3, AKAP2, CDC42BPA, PALM2, DCLK2, NMU, NMUR1, NMUR2 4. COL4A2, FLT1, MMP9, TGFB3, MTDH, DIAPH3, PALM2, DCLK2, NMU, NMUR1, NMUR2 5. COL4A2, FLT1, MMP9, TGFB3,, DIAPH3, PALM2, DCLK2, NMU, NMUR1, NMUR2 1. 6. COL4A2, FLT1, MMP9, TGFB3, MTDH, DIAPH3, PALM2, DCLK2, NMU, NMUR1, NMUR2 IGFBP5, TGFB3, FGF18, ESM1, RARRES3, PITRM1, EXT1, EXTL3, SCUBE2, EBF4,CDC42BPA, CDCA7, CDCA7L, GMPS, MELK, RFC4, WISP1, HRASLS, BBC3, DTL, FBXO31, EGLN1, GNAZ, MTDH, FLT1, ECT2, DIAPH3, NUSAP1, AKAP2, NDC80, PRC1, ORC6L, CENPA, DCK, CCNE2, MCM6, QSOX2, STK32B 7. 6. MMP9, COL4A2 COL4A2, FLT1, MMP9, TGFB3, MTDH, DIAPH3, PALM2, DCLK2, NMU, NMUR1, NMUR2
Chemotherapy benefit in MammaPrint LOW RISK patients (n=252) DDFS: MammaPrint LOW RISK (n=252) BCSS: MammaPrint LOW RISK (n=252) Percent survival 100 80 60 40 20 ET (n=174, 69%) ET+CT (n=78, 31%) HR 0.26 (0.03-2.02) p=0.20 99% 93% Percent survival 100 80 60 40 20 ET (n=174, 69%) ET+CT (n=78, 31%) HR 0.58 (0.07-4.98) p=0.62 99% 97% 0 0 1 2 3 4 5 Time in years 0 0 1 2 3 4 5 Time in years Knauer et al., Breast Cancer Res Treat, 2010 Feb
Percent survival Chemotherapy benefit in MammaPrint HIGH RISK patients (n=289) 100 80 60 40 20 DDFS: MammaPrint HIGH RISK (n=289) ET (n=141, 49%) ET+CT (n=148, 51%) HR 0.35 (0.17-0.71) p<0.01 0 0 1 2 3 4 5 Time in years 88% 76% Percent survival 100 80 60 40 20 BCSS: MammaPrint HIGH RISK (n=289) ET (n=141, 49%) ET+CT (n=148, 51%) HR 0.21 (0.07-0.59) p<0.01 0 0 1 2 3 4 5 Time in years 94% 81% 12% absolute benefit 50% relative benefit 13% absolute benefit 68% relative benefit Knauer et al., Breast Cancer Res Treat, 2010 Feb
MammaPrint in LN+ patients S. Mook et al. (2009) Breast Cancer Res Treat.
What are Intrinsic Molecular Subtypes? Molecular subtypes show which pathway drives cancer growth. Luminal is driven by the estrogen pathway ERBB2 is driven by the HER2 pathway Basal is driven by neither one of them
ERBB2 amplicon cluster Novel unknown cluster Basal epithelial cluster Normal breast like cluster Luminal epithelial gene cluster Sorlie T et al. PNAS. 2001;98-10869-10874
80-gene BluePrint Profile Breast Cancer Analysis of Entire Human Genome ~25,000 Genes Basal, Luminal, and HER2-type BluePrint Basal-Type Luminal-Type classification HER2-Type Prognostic & Predictive Breast Cancer Genes Identified
IHC and Intrinsic Subtype Not Always Correlated (Gluck et al, Int J Ca, 2013)
22% of HER2+ patients is MammaPrint Low Risk HER2 gene is not part of the 70 genes Distant disease free survival without any adjuvant thereapy 25 143 Knauer et al. Br J Cancer, 2010
Intrinsic Subtype Clinical Assay Development Took the initial instrinsic classifer of 496 genes Developed a final classifier consisting of 50 genes and 5 centroids (provided at https://genome.unc.edu/). The CV classification accuracy of the 50 genes versus the 496 genes was 93%. The assay is called the PAM50 Parker et al., JCO, February 9, 2009
PAM50 by NanoString ncounter Extract RNA from FFPE tumor sample Run RNA & PAM50 CodeSet on ncounter Analysis System Capture patient expression profile Determine Intrinsic Subtype through Pearson s Correlation to Centroids Patient expression profile LumA PAM50 centroids Calculate Risk of Recurrence (ROR) Score ROR =ar LumA + br LumB + cr Her2e + dr Basal + ep+ Pearson s correlation to centroids* Proliferation score (19 genes) ft Tumor size
TransATAC: Study Design ATAC Study Postmenopausal women with invasive BC TAM alone (n = 3,116) TAM + Arimidex (n = 3,125) Arimidex alone (n = 3,125) TransATAC Study (N = 1,017) Samples from AstraZeneca -sponsored ATAC study of anastrozole (aromatase inhibitor) 2008 21 gene test study: RS valid in patients treated with aromatase inhibitors 2011 PAM50 study: Tested excess RNA from 2008 study with PAM50 Dowsett M, et al. J Clin Oncol. 2013;31(22):2783-2790.
TransATAC: ROR Score Discriminates Recurrence Risk Within Nodal Subgroups Percent Without Distant Recurrence 1.0 0.9 0.8 0.7 0.6 0.5 0.4 Node-Negative Patients Low (n=431) Medium (n=180) High (n=128) 2 4 6 8 10 Follow-up Time, years Percent Without Distant Recurrence 1.0 0.9 0.8 0.7 0.6 0.5 0.4 Node-Positive Patients (1-3 nodes) Low (n=6) Medium (n=74) High (n=134) 2 4 6 8 10 Follow-up Time, years Risk Group N (%) Events a % Without Recurrence at 10 years Low 431 (58) 17 96% (94%-98%) Intermediate 180 (24) 22 86% (81%-92%) High 128 (17) 38 67% (59%-76%) a Number of events through 10 years. 1. Dowsett, M. et al. J Clin Oncol. 2013;31(22):2783-2790. Risk Group N (%) Events a % Without Recurrence at 10 years Low 6 (3) 0 100% (N/A) Intermediate 74 (35) 11 84% (76%-93%) High 134 (63) 38 68% (59%-77%) a Number of events through 10 years. 32
Prosigna (PAM50) Gene Signature Assay Risk Groups Better Characterize Late Recurrence Prosigna provides greater insight into probability of breast cancer recurrence between Year 5 and 10 after diagnosis Beyond 5 years, the ROR score discriminates the risk of distant recurrence much better than RS Percent without Distant Recurrence, % 100 95 90 85 80 Low ROR score High ROR score Low RS score High RS score 80 100 95 90 85 RS=Recurrence score 0 2 4 6 8 10 Follow-up Time, years Sestak I, et al. J Natl Cancer Inst. 2013;105(19):1504-1511.
ABCSG-8: Study Background and Patients Included in PAM50 Analysis Primary Surgery 3,714 eligible for ABCSG-8 Randomization 1,671 re-consented or deceased Tamoxifen 20 mg 2 years Tamoxifen 20 mg 3 years Tamoxifen 20 mg 2 years Anastrozole 1 mg 3 years 1,620 FFPE blocks collected 25 insufficient tumor tissue in specimen 73 insufficient RNA isolated 44 failed QC specs for device 1,478 evaluable tissue specimens Gnant M, et al. Ann Oncol. 2014;25(2):339-345.
ABCSG8 Study: Prosigna Identified Low-Risk Patients Node negative Patients Node positive Patients (1-3 nodes) Risk Group N (%) Events % without recurrence at 10 yr Low 487 (47%) 15 97% [94%-98%] Intermediate 335 (32%) 28 90% [86%-93%] High 225 (21%) 32 84% [78%-89%] Total 1,047 Risk Group N (%) Events % without recurrence at 10 yr Low 158 (41%) 7 94% [88%-97%] High 224 (59%) 46 76% [69%-81%] Total 382 Gnant M, et al. Ann Oncol. 2014;25(2):339-345.
ABCSG8: 15-year risk of distance recurrence Risk group Number of patients (%) Number of Events through 10-years Low 502 (34%) 15 Estimated % Without Recurrence at 10 years [95% CI] 15 years [95% CI] 96.7% [94.6%-98.0%] 95.6% [92.7%-97.3%] Intermediate 478 (32%) 35 High 498 (34%) 87 91.3% [88.1%-93.8%] 79.9% [75.7%-83.4%] 87.3% [82.3%-90.9%] 72.1% [65.2%-77.8%] Total 1,478 (100%) 137 89.5% [87.7%-91.1%] 84.9% [81.9%-87.5%] 36 Gnant M et al. Ann Oncol. 2014;25(2):339-345. Gnant M et al. Ann Oncol 2013 May 1, 2013;24(suppl 3):iii29.
Danish Breast Cancer Group (DBCG) study of endocrine therapy DBCG is the national cooperative group in Denmark Sets treatment guidelines for all patients in Denmark strictly followed Collects outcome data for all patients in Denmark into a single centralized, registration-quality database Facilitates collection of tumor blocks From 1999-2003 all high risk HR+, post-menopausal patients received 5 years of endocrine therapy alone Node-positive or Node-negative with T>2cm, or Grade II/III First genomic study of a comprehensive national cohort of breast cancer patients 2971 patients eligible in the database in this period 2749 blocks collected (92%) Node positive : N = 1480 Late recurrence analysis: N = 2722 Analyzed ROR and subtype ability to predict 10 year recurrence, late recurrence, and recurrence for 1, 2, 3+ nodes (risk cutpoints depending on #nodes)
DBCG 10 year Distant Recurrence Analysis* Cumulative Incidence by Risk Group Cumulative Incidence by Subtype All Patients * P values Nodal Status Risk Category 10-Year DR [95% CI] Any Diff. Diff. from Int Intermediate 9.6 [7.4-12.2] <0.0001 High 20.8 [18.3-23.4] <0.0001 Low 4.3 [2.9-6.2] 0.0005 High 18.5 [14.9-22.4] <0.0001 Node- Intermediate 7.3 [4.8-10.5] <0.0001 Negative Low 4.9 [2.8-7.8] 0.1543 Node- High 21.9 [18.9-25.1] Positive (1-3 <0.0001 NA nodes) ** Low 4.8 [3.1-6.9] Intrinsic Subtype Node Negative N [%] Node-Positive (1-3 Nodes) Prob of 10-yr DR [95% CI] Node-Negative Node-Positive (1-3 Nodes) Luminal A 632 [50.3%] 883 [60.2%] 6.3% [4.4-8.6] 8.7% [6.7-10.9] Luminal B 502 [40.0%] 475 [32.4%] 14.1% [10.9-17.7] 22.2 [18.1-26.7] Luminal A/B P-value - - <0.0001 <0.0001 Laenkholm et al, ASCO poster 2015
DBCG Late Distant Recurrence Cumulative Incidence by Risk Group Cumulative Incidence by Subtype Patient Risk Prob of late DR P values N (%) Group [95%CI] Any Diff. Diff. from Int. High 870 (40%) 10.2% [8.0-12.7] 0.0091 Intermediate 650 (30%) 6.1% [4.2-8.6] <0.0001 Low 644 (30%) 2.4% [1.3-4.1] 0.0074 Subtype N (%) Prob of late DR [95%CI] Any Diff. Diff. from LumA Luminal A 1281 (59%) 4.5% [3.3-5.9] - Luminal B 733 (34%) 10.3% [7.8-13.1] 0.0002 <0.0001 Her2-Enriched 132 (6%) 8.8% [4.4-15.0] 0.0347 Basal-Like 18 (1%) - - - Laenkholm et al. Poster ASCO 2015
DBCG Comprehensive Cohort Study: Analysis of Node Positive Patients Cumulative Incidence for 1 Positive Node Cumulative Incidence for 2 Positive Nodes Characteristic Number of Positive Nodes 1 2 3 Age N (% of 1+node) N (% of 2+node) N (% of 3+node) 50-59 317 (39%) 172 (40%) 93 (40%) 60-69 328 (41%) 166 (39%) 95 (41%) 70 164 (20%) 88 (21%) 43 (19%) Tumor Size 10 81 (10%) 32 (8%) 11 (5%) 11-20 415 (51%) 185 (43%) 94 (41%) 21-30 209 (26%) 136 (32%) 76 (33%) >30 104 (13%) 73 (17%) 50 (22%) Histological Subtype Ductal 663 (82%) 352 (83%) 191 (83%) Lobular 114 (14%) 62 (15%) 34 (15%) Other 32 (4%) 12 (3%) 6 (3%) Histological Grade 1 274 (34%) 129 (30%) 77 (33%) 2 364 (45%) 198 (46%) 101 (44%) Nodal Status Risk Category N 10-Year DR [95% CI] All Patients (1-3 Positive Nodes) 1-Positive Node 2-Positive Nodes 3-Positive Nodes High 703 22.4% [19.1-25.9] P values Any Diff. Diff. from Int <0.0001 Intermediate 400 11.8% [8.4-15.9] <0.0001 - Low 363 3.8% [2.0-6.3] 0.0007 High 274 21.0% [15.9-26.6] 0.0202 Int 237 14.9% [9.9-20.9] <0.0001 - Low 298 3.6% [1.7-6.5] 0.0001 High 224 20.7% [15.2-26.8] 0.0034 Int 137 8.7% [4.4-15.0] 0.0007 - Low 65 4.6% [1.2-11.8] 0.4165 High 205 26.1% [19.5-33.0] 0.0086 Int 26 0% [NA] 0.0086 - Low N/A - - 3 85 (11%) 52 (12%) 28 (12%) Source: Ejlertsen et. al., ASCO 2015, Poster #513 Not done 86 (11%) 47 (11%) 25 (11%) 21
The Future
Prospective Validation of Oncotype DX: The TAILORX Trial 11,248 ER+/LN- patients Low RS: Hormonal Therapy High RS: Chemo + Hormonal Therapy Hormonal Therapy Chemo + Hormonal Dowsett, M. & Dunbier, A. Clin Cancer Res, 2008.
Kaplan Meier Estimates in the Analyses of Invasive Disease free Survival, Freedom from Recurrence of Breast Cancer at a Distant Site, Freedom from Recurrence at Any Site, and Overall Survival. Sparano JA et al. N Engl J Med 2015;373:2005-2014.
Prospective Validation of Oncotype DX for N(+): RxPonder (S1007)
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Piccart, AACR, 2016
Implications for Clinical Practice MINDACT is clearly practice changing Genomic high/clinical high has 91% DMFS with modern screening and chemotherapy Genomic low/clinical high has 5 years DMFS of 95% when treated with ET alone Includes 1-3 positive node Includes ER positive, Her2 positive (node neg?) Chemo saved in 48% of clinical high risk patients
Comparison of Gene Sets Only 34 of these genes are known to be involved in a specific pathway* *According to http://www.genome.jp/kegg/mapper.html
NCCN Guidelines Update 1.2016
What Do We Do Now? We should attempt some sort of prescreen (clinicalpathologic variables, Adjuvant! Online, NHS Predict, Magee Equations) prior to ordering multiparametric genomic test If insurers allow ONE test ER (+) LN(-) Mammaprint, Pam 50 or ODx if Magee Equation Int/High Mammaprint in Her2 (+), T1, ER pos ER (+), LN (+): Mammaprint, PAM50, or ODx in postmeno LN (+) Mammaprint in premeno LN (+)