Tumorbiologische Aspekte effektive prädiktive Diagnostik aus pathologischer Sicht

Transkript

1 Tumorbiologische Aspekte effektive prädiktive Diagnostik aus pathologischer Sicht Carsten Denkert Translational Cancer Research Group Institute of Pathology Charité Universitätsmedizin Berlin Berlin, Germany AGO, , Kassel

2 Conflict of interest statement Research funding, honoraria, shareholder: Sividon Diagnostics Research funding: Siemens Medical Solutions

3 Übersicht Tumorbiologie beim Mammakarzinom St. Gallen 2011 ER, PR, HER2, Ki67 Genexpressionstests in der Pathologie

4 E _G H&E Estrogen Receptor Progest. Receptor HER2 HER2 SISH DISKUS 027

5 E _G ER H&E Estrogen Receptor Progest. Receptor HER2 HER2 SISH DISKUS 030

6 E _G PR H&E Estrogen Receptor Progest. Receptor HER2 HER2 SISH DISKUS 032

7 E _G H&E Estrogen Receptor Progest. Receptor HER2 HER2 SISH DISKUS 028

8 E _G HER2 H&E Estrogen Receptor Progest. Receptor HER2 HER2 SISH DISKUS 036

9 Ki67 ER HER2 Endocrine therapy alone Endocrine ± cytotoxic therapy Cytotoxics + anti HER2 + endocrine therapy Cytotoxics + anti HER2 therapy Cytotoxic therapy adapted from Goldhirsch 2011

10 Grenzen der Immunhistochemie Ki67 ER HER2 Technische Fragen Endocrine therapy Quantifizierung von Markers alone und cutoffs e.g. Ki67 Endocrine ± Interobserver Variabilität cytotoxic therapy Assay Standardisierung Cytotoxics + anti HER2 + endocrine therapy Prädiktive Fragestellungen: Endocrine Tx vs. Chemoendocrine Tx Cytotoxics + anti Verschiedene Typen HER2 von anti therapy HER2 Therapy Triple negative Karzinome Cytotoxic therapy

11 HER2 beim Mammakarzinom

12 Zwei Säulen der HER2 Diagnostik Immunhistologie 0, In situ Hybridisierung (FISH, SISH, CISH) Ratio HER2 vs Chr 17 Nicht amplifiziert (<1.8) Equivocal ( ) Amplifiziert (>2.2)

13 Standardisierte Scoring Systeme HER 2 ( DAKO Score ) modified 2007 ASCO CAP Zellzahl Intensität Membran Score Beurteilung <10% 0 negativ >10% partiell 1+ negativ >10% schwach komplett 2+ schw. pos % stark komplett 2+ schw. pos. >30% stark komplett 3+ stark positiv American Society of Clinical Oncology/College of American Pathologists guideline recommendations JCO 2007

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15 HER2 SISH DISKUS 001

16 ASCO CAP guidelines update 2011 Was macht man im Grenzbereich? Immuno Grenzbereich: 10-30% stark ISH Grenzbereich: Als Pathologe: mit der jeweils anderen Methode überprüfen Als Kliniker: Trastuzumab Therapie nach FDA Kriterien ab >10% starke Expression ab >2.0 Ratio in der ISH

17 ASCO CAP Guidelines update 2011 Als Pathologe: mit der jeweils anderen Methode überprüfen

18 ASCO CAP guidelines update 2011 Was macht man im Grenzbereich? Immuno Grenzbereich: 10-30% stark ISH Grenzbereich: Als Pathologe: mit der jeweils anderen Methode überprüfen Als Kliniker: Trastuzumab Therapie nach FDA Kriterien ab >10% starke Expression ab >2.0 Ratio in der ISH

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20 Fallbeispiel: IHC 20% starke Färbung, FISH negativ Klinische Konsequenz?

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22 GeparSixto Studie zentrale Pathologie Prospektive zentrale Pathologie vor Randomisierung ER, PR, HER2 Immuno + SISH Ki67 Dokumentation in Medcodes Befundbericht an Zentrum (und lokalen Pathologen)

23 Limitations of immunohistochemistry Comparison of central and local IHC in clinical trials of the GBG False positive rate in local HER2 testing GeparQuattro N=217 local HER GeparSixto* N=158 local HER Preliminary

24 Higher response rates in centrally HER2 positive cases regardless of method used pcr: ypto/is ypn0 Denkert et al., 2013

25 GeparTrio n=278, Noske et al,. BRCT 2011 % HER2 positive cases Concordance

26 Wie % der Fälle sind HER2 positiv? HER2 Monitor, Choritz, Kreipe et al, 2011 HER2 pos Rate: 16.7% (95% CI %) HER2 Bestimmungen 42 Pathologien, davon 6 Pathologien = outlier 36 Pathologien berücksichtigt

27 Ergebnis der Ringversuche für die HER2-IHC von Wasielewski, Virchows Archiv 2008

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29 HER2 IH

30 HER2 SISH

31 GeparQuattro: HER2+ (IHC/ISH) cases with neg HER2mRNA: they exist, but they have a low pcr rate All cases including centr HER2 neg Only centr HER2 positive cases Denkert et al., 2013

32 Ki67 beim Mammakarzinom

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35 Best cutoff: 13,25% N=144 N=127 JNCI, 2009, 101:

36 SABCS 2012: Ergebnisse des internationalen Ringversuches

37 2011 Review: Ki67 1. Ki67: independent prognostic factor for diseasefree survival (LoE1b (Simon et al)). 2. Ki67: not predictive for long term follow up after chemotherapy 3. Ki67: associated with pcr in the neoadjuvant setting (LoE 2b) 4. Standardization of scoring methods and techniques is needed.

38 Core biopsy: uni/bilateral ct2 4a d cn0 3 size 2 cm* *low risk patients were excluded (T2 + ER/PR pos. + cno + G1/2 + > 35 yrs) NC Sonography CR/ PR R R OP OP OP NX TACx6 TACx6 OP TACx8 von Minckwitz et al, JNCI 100: 542, 2008 von Minckwitz et al. JNCI 100; 552, 2008 Study cohort GeparTrio n=1166 This presentation is the intellectual property of the author/presenter. Contact them at carsten.denkert@charite.de for permission to reprint and/or distribute.

39 pcr all cases p< Ki67 15% vs % vs. >35% DFS all cases p< OS all cases p< pcr rate (%) disease free survival rate overall survival rate Ki67 15% Ki % Ki67>35% time (years) time (years) Multivariate analysis: p< p=0.012 p=0.013 This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

40 Systematic cutpoint analysis for Ki67 pathological complete response (pcr) after neoadjuvant chemotherapy Odds ratio with 95% CI 15% 35% pcr Ki67 (% positive cells) This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

41 Systematic cutpoint analysis for Ki67 pathological complete response (pcr) after neoadjuvant chemotherapy Odds ratio with 95% CI 15% 35% 93 of 94 cutpoints significant pcr Ki67 (% positive cells) This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

42 Evaluation of different molecular subtypes Molecular subtype radar diagram (MSRD) HR positive HR+/HER2 HR+/HER2+ HER2 negative HER2 positive HR /HER2 HR /HER2+ HR negative This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

43 Evaluation of different molecular subtypes Molecular subtype radar diagram (MSRD) HR positive HR+/HER2 HR+/HER2+ HER2 negative HER2 positive HR /HER2 HR /HER2+ HR negative This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

44 Ki67 15% pcr rate (%) HR positive HR+/HER2 HR+/HER2+ HER2 negative HER2 positive HR /HER2 HR /HER2+ HR negative This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

45 Ki67 intermediate pcr rate (%) HR positive HR+/HER2 HR+/HER2+ HER2 negative HER2 positive 15% 15 35% HR /HER2 HR /HER2+ HR negative This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

46 Ki67 >35% pcr rate (%) HR positive p< HR+/HER2 p< HR+/HER2+ p=ns HER2 negative p< % HER2 positive p= % HR /HER2 p=0.003 >35% HR negative p< HR /HER2+ p=ns This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

47 Overall survival HR positive HR+/HER2 HR+/HER2+ HER2 negative HER2 positive HR /HER2 HR /HER2+ HR negative This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

48 Ki67 15% Overall survival (mean OS months) HR positive HR+/HER2 15% HR+/HER2+ HER2 negative HER2 positive HR /HER2 HR /HER2+ HR negative This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

49 Ki67 intermed. Overall survival (mean OS months) HR positive HR+/HER2 15% HR+/HER % HER2 negative HER2 positive HR /HER2 HR negative HR /HER2+ This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

50 Ki67 >35% Overall survival (mean OS months) HR positive p< HR+/HER2 p= % 15 35% HR+/HER2+ p=ns HER2 negative p=0.001 HER2 positive p=ns >35% HR /HER2 p=ns HR negative p=ns HR /HER2+ p=ns DFS: similar results (not shown) This presentation is the intellectual property of the author/presenter. Contact them at for permission to reprint and/or distribute.

51 Ki67 combination of predictive and prognostic effects Ki67: negative prognostic marker and positive predictive marker. Positive predictive effect: in 6 of 8 subtypes (increased pcr). Negative prognostic effect: only in HR+ subtypes where the response rates are low and where pcr is not prognostic. Tumor types, type of therapy and response rates will have influence on the observed cutpoints for Ki67. This presentation is the intellectual property of the author/presenter. Contact them at carsten.denkert@charite.de for permission to reprint and/or distribute.

52 Ki67 in GeparTrio parallel Investigation of core biopsy and surgical specimen 1166 cores, 667 surgical tissues Standardized counting of 200 cells Large sections (no TMA) End points: pcr (prediction) DFS, OS (prognosis)

53 Ki67 in residual tumor after NACT GeparTrio n=1151 pcr 15,1 35% <15% >35% von Minckwitz, ASCO 2012

54 Subgroups: Ki67 after NACT GeparTrio HR positive, n=691 HR negative, n=460 pcr <15% pcr >35% 15,1 35% <15% 15,1 35% >35% von Minckwitz, ASCO 2012

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57 total cells positive cells

58 ??????????????????????????????????????????

59 Workflow for Ki67 analysis Select target area (20x Obj.) Digital image on computer monitor Copy paste image to Ki67 software (local or web based) Identify cells, count positive and negative cells Check for typical software errors and correct them Report Ki67 value

60 Select target area (20x Obj.) Digital image on computer monitor Copy paste image to Ki67 software (local or web based) Identify cells, count positive and negative cells Check for typical software errors and correct them Report Ki67 value

61 Select target area (20x Obj.) Digital image on computer monitor Copy paste image to Ki67 software (local or web based) Identify cells, count positive and negative cells Check for typical software errors and correct them Report Ki67 value

62 Select target area (20x Obj.) Digital image on computer monitor Copy paste image to Ki67 software (local or web based) Identify cells, count positive and negative cells Check for typical software errors and correct them Report Ki67 value

63 To do list for validated image analysis Develop a userfriendly system Validate in clinical study Document error rate of typical errors Validate in multicenter setting Frederick Klauschen and Stephan Wienert

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66 Ki67 Zusammenfassung Einfacher, in jeder Pathologie durchführbarer Marker, kostengünstig. Viele cutpoints sind biologisch sinnvoll. Interobserver Variabilität, keine international geltenden Cutpoints. Wichtig: Lokale Validierung des Tests. Keine validen Aussagen im Grenzbereich! Zukunft: Bildanalyseverfahren

67 Hormonrezeptoren beim Mammakarzinom

68 Wann ist ein Tumor HR positiv? ASCO/CAP 2010: geänderter cutoff für Hormonrezeptoren 1% = positiv (früher 10%) Harvey, 1999, Concordance IHC and LBA 85.5% Hammond, 2010

69 Schwach ER pos Tumoren haben oft einen basalen Phänotyp Iwamoto, JCO, 2012

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71 Neue Möglichkeiten der Genexpressionsanalyse

72 Ki67 ER HER2 Endocrine therapy alone Endocrine ± cytotoxic therapy Cytotoxics + anti HER2 + endocrine therapy Cytotoxics + anti HER2 therapy Cytotoxic therapy adapted from Goldhirsch 2011

73 Hormonrezeptor positive Karzinome 70% Wann hat eine Patientin mit HR+ und Her2 neg. Mammakarzinom mit alleiniger endokriner Therapie eine sehr gute Prognose? Anti Hormontherapie ± Chemotherapie Möglichkeiten: 1. Klinisch pathologisch 2. EndoPredict 3. Oncotype Dx 4. PAM50 5. Mammaprint 6. upa PAI1

74 Recurrence Score Oncotype DX 16 genes and 5 control genes Sparano JCO 2007

75 Recurrence Score Oncotype DX 16 genes and 5 control genes Sparano JCO 2007

76 EP score und EPclin score Member 1 BIRC5 RBBP8 Reference genes CALM2 OAZ1 RPL37A Member 2 Member 3 UBE2C IL6ST AZGP1 DHCR7 EP Score + Tumorgröße + Nodalstatus Member 4 MGP STC2 EPclin Score Filipits et al. Clinical Cancer Res. 2011

77 EndoPredict Test

78 EndoPredict-assay Decentral testing in clinical pathology laboratories RNA Isolation EndoPredict Test Evaluation FFPE-Slide RNA Isolation: Manuell (Siemens) Automatisiert (Siemens) RT qpcr: Evaluation 3h, 12 Proben 4h, 48 Proben 2h pro Platte 15 min time < 8 h

79 Development of the Endopredict assay Training Validation I Validation II EP ABCSG 6 ABCSG 8 n = 378 n = Filipits et al. Clinical Cancer Res. 2011

80 EPclin Score in subgroups Nodal status pn0 vs.pn1 Luminal A und B (Ki67) Grading G1 vs G2 vs G3 Filipits et al. Clinical Cancer Res. 2011

81 Prognostic performance of markers Nodal Nodal + size Nodal + size + age + grade + ER + Ki67 Nodal + size + age + grade + ER + Ki67+ EP Adjuvant Adjuvant + EP Adjuvant + ER + Ki67 Adjuvant + ER + Ki67 + EP P=0.039 P=0.024 P < EPclin ABCSG c index 0.82 Filipits et al. Clinical Cancer Res. 2011

82 Die Entwicklung des Endopredict Tests

83 Übersicht S e it e 83 AGO Breast Committee. Diagnosis and Treatment of Patients with Primary and Metastatic Breast Cancer. Recommendations online.de

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86 Simon JNCI, 2009

87 Simon JNCI, 2009

88 Annals of Oncology, 2012

89 Level of Evidence (Simon et al 2009) Annals of Oncology, 2012

90 Vergleich Oncotype Dx vs. EndoPredict Oncotype Dx quant. RT PCR, 16 Gene FFPE Gewebe reiner Gen Score low/intermediate/high enthält u.a. ER und HER2 IHC bietet ähnliche Info Zentrallabor Kalifornien Keine ärztliche Leistung in Deutschland ca. 4125$ /3100 EndoPredict quant. RT PCR, 8 Gene FFPE Gewebe integriert pt und pn Status Keine intermediäre Risikogruppe ohne ER bzw. HER2 unabhängige Information lokal in Patho durchführbar, im Ringversuch validiert Ärztliche Leistung im deutschen Gesundheitssystem ca. 2000

91 Molekulare Pathologie: EndoPredict neue Möglichkeiten zur Prädiktion der Response 1. Wann hat eine Patientin mit ER+ und Her2 neg. Mammakarzinom mit alleiniger endokriner Therapie eine sehr gute Prognose? 2. Können dezentrale molekulare Genexpressions Analysen (RT PCR in Paraffingewebe) beim Mammakarzinom diese Frage reproduzierbar beantworten?

92 Development of the Endopredict assay Training Validation I Validation II Method validation Transfer Analytical validation EndoPredict proficiency testing program 2011 in 7 Pathology Institutes Berlin, Gelsenkirchen, Hannover, Heidelberg, Munich, Vienna, Zürich Filipits et al 2011, Bohmann et al. 2009, Müller et al. 2011, Denkert et al. 2010, Denkert et al. 2012

93 EndoPredict 2013 Referenzwert + Teilnehmer Threshold Denkert et al. Virchows Arch

94 Additional testing in a total of 15 laboratories

95 BMC Cancer, 2012 Ann Oncol, 2012

96 Dubsky et al. Ann Oncol, 2012

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98 Endocrine Therapy after Tamoxifen in postmenopausal patients AGO e. V. in der DGGG e.v. sowie in der DKG e.v. Guidelines Breast Version After 5 yrs. tamoxifen (EAT) AI up to 3 to 5 yrs. 1b A ++ Node-positive disease 2b B ++ Long tamoxifen-free interval 2b B + Consider EAT with AI for pts. who changed to postmenopausal status during 5 yrs. Tam Oxford / AGO LoE / GR Continuation of Tam up to total 10 yrs. 1a A ++

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101 Wie oft ändert sich die Therapie? Endopredict Tests im ersten Jahr an der Charité n=167 Angaben zur Therapie für 130 Patientinnen, retrospektiv Berit Müller, 2012

102 Gnant et al, SABCS 2012

103 Gnant et al, SABCS 2012

104 Voraussetzungen für die Durchführung des EndoPredict Tests Klinische Situation: 1. HR positiver, HER2 negativer Tumor 2. Klinisch relevante Frage: Ist eine alleinige endokrine Therapie ausreichend? Organisation: 1. Ein Paraffinblock aus dem Tumor 2. Histobericht mit Angabe von Tumorgröße und Nodalstatus 3. Ü-Schein an die Pathologie zur molekularen Diagnostik Testdurchführung an der Charité*, ca. 2-5 Arbeitstage (*auch an derzeit ca 30 weiteren Pathologischen Instituten in Deutschland möglich)

105 RESPONSIFY FP7 Project (Scientific Coordinator: Sibylle Loibl, 12 European partners) Aim:... to develop FFPE based predictive IVD tests for anti HER2 and antiangiogenic therapies...

106 Charité Britta Beyer Jan Budczies Silvia Darb Esfahani Frederick Klauschen Ines Koch Berit Müller Judith Prinzler Bruno Sinn Petra Wachs Manfred Dietel Siemens Guido Hennig Sividon Kerstin Bohmann Christoph Petry Ralf Kronenwett Karsten Weber Jan Brase ABCSG Martin Filipits Peter Dubsky Michael Gnant Margaretha Rudas GBG Gunter von Minckwitz Sibylle Loibl Valentina Nekljudova Keyur Mehta Yasmin Issa Nummer Christiane Mayr Translationales Subboard Neoadjuvantes Subboard