PD-L1 TestingGerman Experience

P. Schirmacherfor

QuIP

Quality Management Molecular Diagnostics

• Method/Inter-Center-Optimisation/Validation• Preclinical Validation/internal QM • Accreditation Institutes (DAkkS; ISO 9001 DIN

17020)• National Round Robins (QuIP)

QuiP (DGP)• Independent• Self administered• Non commercial• Expert guided (Panels)• Extensive RR-Expertise (> 300 RRs)• High acceptance (diagnostic community, clinicians, industry,

politics)• International (in selected indications)• ESP-LOA and cooperation

Free Choice of Method, e.g.• Sanger• Pyro-Sequencing• Mutation-spezifische PCR• NGS (Deep/Panel

Sequencing)Reflection of technologyEvaluation by diagnostic result

Principle: Free Choice of Methods

Constant Round Robins2010 2011 2012 2013 2014 2015 2016 2017 2018

RfB BRAF 600 BRAF 600 BRAF 600 BRAF 600RfB CD117 CD117 CD117 CD117 CD117 CD117 CD117 CD117RfB EGFR EGFR EGFRRfB Erez/PRez Erez/PRez Erez/PRez Erez/PRez Erez/PRez Erez/PRez Erez/PRezRfB GIST GIST GIST GIST GISTRfB HER2n IHC HER2n IHC HER2n IHC HER2n IHC HER2n IHCRfB HER2n ISH HER2n ISH HER2n ISH HER2n ISH HER2n ISHRfB Keratine Keratine Keratine Keratine Keratine Keratine Keratine KeratineRfB KlonML KlonML KlonML KlonML KlonML KlonML KlonMLRfB KRASRfB Lymphome Lymphome Lymphome Lymphome Lymphome Lymphome Lymphome LymphomeRfB MMRD MMRD MMRD MMRD MMRD MMRD MMRD MMRDRfB MSI MSI MSI MSI MSI MSI MSIRfB NEM NEM NEM NEM NEM NEM NEM NEMRfB RAS RAS RAS RASmultiblock ER ER ER ER ER ER ER ERmultiblock PR PR PR PR PR PR PR PRmultiblock Her2-IM Her2-IM Her2-IM Her2-IM Her2-IM Her2-IM Her2-IM Her2-IMmultiblock Her2-ISH Her2-ISH Her2-ISH Her2-ISH Her2-ISH Her2-ISH Her2-ISH Her2-ISHmultiblock Ki-67 Ki-67 Ki-67 Ki-67 Ki-67 Ki-67 Ki-67 Ki-67provitro HPV HPV HPV HPV HPV HPV HPV HPVprovitro TBc TBc TBc TBc TBc TBc TBc TBcRfB T790M T790MRfB PD-L1MMRfB PD-L1 NSCLC PD-L1 NSCLC

RfBEGFR IHC sqNSCLC

RfB BRCA1/2 BRCA1/2RfB BRAF 600 Lunge

Round Robin Status

T790M 2016 finished

PD-L1 NSCLC final report

EGFR IHC sqNSCLC

final report

PD-L1 Melanoma(2)

final report

T790M 2017 (2) ongoing Q1-Q2

PD-L1 NSCLC (2) ongoing Q1-Q2

BRAF V600 (2) planned Q3-Q4

BRCA 1/2 (2) planned Q3-Q4

Prototypic Round Robins

Round Robin Status

MLH1ConsiderationQuIP-Board

ROS1ConsiderationQuIP-Board

IDH1(R132H) ConsiderationQuIP-Board

Implementation of new Diagnostic Assays

Quality Management• Panel-establishment and -validation (> 3 m)

• Establishment of expert panel (n~5-8; Lead-Institutes; other Panel centers)

• Consent about approach (methodology, time schedule, testing conditions etc.)

• Identification and selection of suitable test cases• Cross validation by lead institutes• Testing/validation of panel centers; evaluation;

set-up of RR conditions

• National/international round robin (> 2 m)• announcement/feed-back; application• preparation by Lead-institutes, RfB• Probe mailing, testing, test results mailed• Evaluation and publication

Information about approval of new drug with predictive molecular testing (1-3 months ahead)

Implementation• Validation and optimisation of testing

modalities• Solutions for open scientific issues• Diagnostic recommendations, information,

discussion with clinical societies• Diagnostics for early accession/

compassionate use programs• Broad/nationwide availability (methodology,

expertise, training)• Agreement on reimbursement issues• Local issues• Others (e.g. legal issues)

Implementation without regulatory, financial and intellectual framework but decisive for drug/therapy success (system relevant).Requires Competence Center System (Peers) and more attention by industry and regulatory authorities

PD-L1 IHCGerman Strategy for Roll-out and QM

• National Harmonisation Study (DGP)– Validation– Harmonisation– Recommendation

• National QA Measures (QuIP)– Quality Assurance– Teaching and Training

• National Improvement Activity (nNGM/DKH)

Pathology: Industrial PartnersReinhard Büttner BMSManfred Dietel MSDLukas Heukamp RocheKorinna Jöhrens AstraZenecaThomas Kirchner VentanaSimone Reu DakoJosef Rüschoff TargosAndreas ScheelHans-Ulrich SchildhausPeter SchirmacherMarkus TiemannArne WarthWilko Weichert

PD-L1 IHC in NSCLCNational Harmonisation-Study

Response to Anti PD-1 (Pembrolizumab) and Expression of PD-L1 in NSCLC

Different Antibodies and Antibody Reactivities

Scheel et al., 2016

Tumorimmunotherapy: PDL-1-Testing

Besse et al, Oral Presentation at ECC 2015

BIRCH: PD-L1 TC and IC Selection Criteria

• BIRCH enrolled patients with tumors that were PD-L1 TC2/3 or IC2/3– VENTANA SP142 IHC assay was used to determine PD-L1 expression on both TC and IC – Archival or freshly collected tumor specimens were required for PD-L1 testing at a

central laboratory

• PD-L1 as a predictive biomarker:– PD-L1 expression on TC and IC was independently predictive of response in patients with previously

treated NSCLC (i.e., POPLAR)1,2

Intrinsic PD-L1 expression in tumor cells (TC)

Adaptive PD-L1 expression in tumor-infiltrating immune

cells (IC)

TC3 or IC3 = TC ≥ 50% or IC ≥ 10% PD-L1+ cells; TC2/3 or IC2/3 = TC or IC ≥ 5% PD-L1+ cells, respectively.1. Horn L, et al. ASCO 2015; 2. Vansteenkiste J, et al. ECC 2015.

IC2/3TC2/3

34% of screened patients between

Jan. 2014 and Dec. 2014

A Global NSCLC-PDL1 Score?

Scheel et al., 2016

Interobserver-Validation (Phase I)

Scheel et al., 2016

Harmonisation Study: Immune Cells

Scheel AH et al., Modern Pathology 2016

Assays:

- Performance in multiple Institutes / Reliability

- Staining pattern - validation of Phase 1; fixed assay

vs. open protocol

- Inter-lab concordance

Evaluation:

- Validation proportional score (6-step)

- Local vs. central evaluation

Phase 2

- Trial analog protocolsDako 22C3, 28-8Ventana SP142, SP263

- Open protocol: 22C3, 28-8(local protocols)

- ≥3 institutes per protocol

TargosMolecular Pathology;

8 PathologyInstitutes

Phase 2

- Project management Phase 2

- NSCLC-TMAs (project specific):- 'TMA-Master' (3DHistotech)- 1.5 mm cores

- Central provision of slides

PD-L1 Testing

+• Robust, broadly available

technology (IHC)• Good antibodies;

alternatives exist• Robust, relatively stable

assay• Consented algorithm

-• Stand-alone assay/ no

methodical convergence• Singular analyses• Algorithm/reporting

complexity; dynamic• Heterogenous expression;

1% threshold problematic• Insufficient refunding

Prototypic Round RobinPD-L1-NSCLC (1)

28.4.16 Decision for RR, Lead- andPanel Institutes6.16 Coordination of Lead Institutes, selection of cases, trilateral validation15.7-26.7. Testing of the Panel Institutes (internal RR)27.7. -29.7. Evaluation of internal RR-results, Lead/Panel-Institute; meeting, decision for open RR15.9 Public PD-L1 training meeting(Charite)End of 9.16 Open RR

Lead Institutes: Berlin, Cologne, GöttingenPanel Institutes: Hannover, Heidelberg, Jena, Munich TU, Munich LMUComposition: 10 cases; 2 pointseach, 18 points necessary forcertificateNo methodical restriction, sectionsprovidedOptional reporting of methodologyOption to challenge and foreducational

Prototypic Round RobinPD-L1-NSCLC (1)

• Participants: 83 (76 D, 4 A, 3 CH)

• Successful: 60

• Success rate: 72%

Composition

< 1%: 3 cases

1%-49%: 2 cases

50-100 %: 5 cases

Diagramm1

836023

gesamt

mit Erfolg teilgenommen

teilgenommen

Sheet1

gesamt83

mit Erfolg teilgenommen60

teilgenommen23

PD-L1-NSCLC Round RobinAntibodies

PD-L1-NSCLC Challenge

Challenged result: 18 Institutes

• 1 Institute post hoc success

• 4 Institutes: Interpretation problems

• 13 Institutes: Staining problems

Prototypic Round RobinMalignant Melanoma (1)

3.8 Panel-Institutes.16 Decision forRR22.8.16 Decision on Lead- and Panel Institutes9/10.16 Coordination of Lead Institutes, Case selection andreciprocal testing14-28.10 Testing of Panel Institutes (internal RR)9/10 Announcement of open RR9.-25.11.16 open RR28.11-2.12. Evaluation of open RR; announcement of results

Lead Institutes: Berlin, Heidelberg, TU MunichPanel Institutes: Hannover, MunichLMU, Cologne, Hamburg, Göttingen, WürzburgComposition: 10 cases; 2 pointseach, 18 points necessary forcertificateNo methodical restriction, sectionsprovidedOptional reporting of methodologyOption to challenge and foreducational

Prototypic Round RobinPD-L1-Melanoma

• Participants: 44

• Successful: 37

• Success rate: 84%

Composition

5%: 5 cases

PD-L1-Melanoma Round RobinAntibodies

PD-L1-NSCLC and MM Round RobinOther Conclusions

• Diverse use of primary antibodies

• 28-8 and SP263 stain stronger

• 22C3 stains rather weaker

• Discrepancies are rather towards lower stainingintensities

• Establishment using tonsil tissue suggested

Future and Ongoing Activities

• PD-L1-NSCLC (2) ongoing (until 15.5.)• PD-L1 Bladder Cancer – in preparation• PD-L1 MM (2) – in discussion

Other/Better Markers!?

• PD-L1 Gene CNV/expression• MSI• Mutational load• Immunophenotyping• Combinations• RESISTANCE MARKERS

PD-L1 Amplifications/Deletions and/or Mutational Load

Increasing number of mutations

n=78n=40

Budczies et al. 2016

WGS/WES?

Panels?

Others?

Immune Cell Infiltration and Response to Conventional and Personalized Treatment

Lasitschka, F, Schirmacher P, Jäger D, Halama N, et al.

Constant Round Robins

RR

Participants 2016

RR

Participants 2016

BRAF 58 ER 186

CD117 66 PR 177

GIST 39 Her2-IM 198

Her2nlHC 88 Her2-ISH 103

Her2nlSH 52 Ki-67 177

RAS 71

Lymphone 71 HPV 7

MMRD 84 TBC 14

MSI 45

Keratine 88

KlonML 29

NEM 69

EGFR 48

PD-L1 Testing�German ExperienceQuality Management Molecular DiagnosticsQuiP (DGP)Diapositiva numero 4Constant Round Robins�Diapositiva numero 6Diapositiva numero 7PD-L1 IHC�German Strategy for Roll-out and QMPD-L1 IHC in NSCLC�National Harmonisation-StudyDiapositiva numero 10Different Antibodies and Antibody ReactivitiesTumorimmunotherapy: PDL-1-TestingDiapositiva numero 13A Global NSCLC-PDL1 Score?Interobserver-Validation (Phase I)Diapositiva numero 16Diapositiva numero 17Diapositiva numero 18PD-L1 TestingPrototypic Round Robin�PD-L1-NSCLC (1)�Prototypic Round Robin�PD-L1-NSCLC (1)�PD-L1-NSCLC Round Robin�AntibodiesPD-L1-NSCLC �ChallengePrototypic Round Robin�Malignant Melanoma (1)�Prototypic Round Robin�PD-L1-MelanomaPD-L1-Melanoma Round Robin�AntibodiesPD-L1-NSCLC and MM Round Robin�Other ConclusionsFuture and Ongoing ActivitiesOther/Better Markers!?PD-L1 Amplifications/Deletions and/or Mutational LoadImmune Cell Infiltration and Response to Conventional and Personalized TreatmentDiapositiva numero 32Diapositiva numero 33Constant Round Robins�