1. Biomarker of Immunotherapy for Future Lung Cancer Treatment
May 6, 2017 TJCC
Teh-Ying Chou, MD, PhD, MBA

2. Evolution of Lung Cancer Classification –
Molecular Pathology
2000
2008
targets today (2015)
future targets
Non-small cell lung cancer
Adenocarcinoma
Large-cell carcinoma
Squamous cell carcinoma
Small cell lung cancer
EGFR
ALK
ROS-1
KRAS and others
BRAF
HER2
RET
DDR2 & others
MET
FGFR1
PI3K
Molecular Pathology-based Personalized Medicine of Lung Cancer
Small-cell lung cancer
Adenocarcinoma
Adenocarcinoma with
actionable mutations
Large-cell carcinoma
Small-cell carcinoma
Squamous cell carcinoma without oncogenic alteration
Squamous cell carcinoma with
oncogenic alteration
The colors denote different histological subtypes
Lancet 2013; 382:

3. Molecular Pathology-based Precision Medicine of Lung Cancer
lung cancer pathology
1999
2004
2015
Molecular Pathology-based Precision Medicine of Lung Cancer
Histology
Targeted Therapy
Immunotherapy
Precision Medicine

4. Biomarker of Immunotherapy for Future Lung Cancer Treatment
Immunotherapy for Lung Cancer
Biomarkers of Immunotherapy for Lung Cancer
PD-L1 IHC: Prediction Biomarker for Anti-PD-1/PD-L1 Immune Checkpoint Inhibitor Therapy
PD-L1 IHC: The Blueprint Project
PD-L1 IHC: The IASLC Atlas
PD-L1 IHC: The Future Perspectives

5. Immunotherapy

6. Immune Checkpoint Inhibition

12. PD-L1 immunohistochemistry as a biomarker
Presented By Keith Kerr at 2015 ASCO Annual Meeting

13. Presented By Justin Gainor at 2015 ASCO Annual Meeting
Slide 11
Presented By Justin Gainor at 2015 ASCO Annual Meeting

14. Presented By Justin Gainor at 2015 ASCO Annual Meeting
Slide 14
Presented By Justin Gainor at 2015 ASCO Annual Meeting

17. PD-L1 expression correlates with efficacy
Predictive value?
Atezolizumab
Nivolumab
Pembrolizumab
Detection antibody*
SP142
28-8
22C3
IHC platform
Ventana
Dako
Tested cells
NSCLC (IC and TC)
UC (IC)
Lung (TC)
NSCLC (TC)
UC (TC and stroma)
Estimated PD-L1 prevalence in NSCLC
PD-L1+ as ≥5% of TCs
PD-L1+ as ≥50% of TCs
TC1/2/3 & IC1/2/3‡
26%
IC1/2/3 & TC0*
30%
TC1/2/3 & IC0*
11%
~25%1
The next question is around the role of the biomarker, and PDL1 selection…
-clearly this is complex subject, as represented by differences in assays etc
-in short, there 3 different assays, each using different detection Ab, IHC platforms, and the type of cells being measured
Our assay is unique as it measures PDL1 on both TC and IC infiltrates within tumor microenvironment- which we feel is salient to capture the full picture as some tumors express PDL1 on IC (ie bladder), or TC or a mixture of both (as we see with lung)
-BMS measures only TC, and Merck measures TC lung and stromal cells for UBC
-with all this said, there are also unique cutoffs and definition of “positive” within studies, which would rationally lead to differences in resulting prevalence between assays
Verdict is out on which approach is best, and exercise called Blueprint, is tasked to detect level of concordance among tests- goal for Phase 1 readout early 2016.
On the data front- as difficult it is to compare ITT, it is nearly impossible compare subgroups based on a “medley” of differences. But, what is encouraging is that the entire class is showing an enrichment trends (we saw this with Pembro as well as nivo in chekmate 57)…
With our CDx, we clearly see enrichment within BIRCH and POPLAR as Ilze pointed out. Great to help define those patients that might benefit most.
~46%1
PD-L1 expression correlates with efficacy
*TC3 or IC3 = TC ≥ 50% or IC ≥ 10% PD-L1+; TC2/3 or IC2/3 = TC or IC ≥ 5% PD-L1+; TC1/2/3 or IC1/2/3 = TC or IC ≥ 1% PD-L1+; TC0 and IC0 = TC and IC < 1% PD-L1+, respectively.
‡POPLAR (Spira, et al. ASCO 2015); Study-003 (Brahmer, et al. ASCO 2014; KEYNOTE-001 (Garon, et al. AACR 2015)
1. Kerr KM, et al. J Thorac Oncol 2015

18. Mutational Load and Immunotherapy
Presented By Justin Gainor at 2015 ASCO Annual Meeting

19. Mutation Load and Immunotherapy
Mutational Load and Immunotherapy
Mutation Load and Immunotherapy
Presented By Christine Walko at 2016 ASCO Annual Meeting

20. Presented By Padmanee Sharma at 2016 ASCO Annual Meeting
Slide 17
Presented By Padmanee Sharma at 2016 ASCO Annual Meeting

21. Presented By Padmanee Sharma at 2016 ASCO Annual Meeting
Overlap between responders and <br />non-responders make it difficult to use mutational load as a predictive biomarker
Presented By Padmanee Sharma at 2016 ASCO Annual Meeting

22. Highest Mutation Burden Subset Displays Marked Improvement in OS
Presented By Alexandra Snyder Charen at 2016 ASCO Annual Meeting

23. Why Does Mutational Load Matter?
Presented By Alexandra Snyder Charen at 2016 ASCO Annual Meeting

24. Issues with the biomarkers
Heterogeneity- multiple tumors and multiple passes within a tumor
Interval between biopsy and treatment
Primary versus metastatic disease
Antibody and staining conditions
Defining a positive results (cut-offs):
Cell type expressing PD-L1 (immune cell vs. tumor or both)
Location of expression- cell surface vs. intracellular vs. stromal
Intensity, percent of cells “positive”
Distribution- patchy vs. diffuse, intratumoral vs. peripheral

25. “Companion/Complementary Testing” is the gold standard basis of therapy selection, while mutation load and neoantigen measurement etc. are “surrogate testing” as predictive biomarkers

26. The Blueprint Project:
Comparing PD-L1 IHC Diagnostics for Immune Checkpoint Inhibitors

38. Different Classes of In Vitro Diagnostics

39. Blueprint Project: Study Design
Two-phase study to gain sufficient data and rigor:
Phase 1
Feasibility on small cohort evaluated at Dako and Ventana
To assess the four PD-L1 diagnostics on the same NSCLC cases and gather initial data in two stages:
Step 1: Evaluate analytical comparability by quantifying and comparing PD-L1 expression on tumour and immune cells
Step 2: Clinical agreement was assessed through comparisons of patient classification and agreement using various combinations of assays and PD-L1 staining thresholds
Phase 2
TBD: Proposed larger, statistically powered study that will be designed from the Phase 1 “information gathering”

40. Blueprint Project: Analytical Comparison of PD-L1 Staining
28-8
SP263
SP142
Three assays (22C3, 28-8, SP263) demonstrate similar analytical performance with respect to percentages of tumour cells positive and dynamic range
SP142 consistently labels fewer tumour cells
Tumour Cells
22C3
28-8
SP263
SP142
Immune Cells
All assays labeled immune cells, but there was increased analytical variability across assays and readers with immune-cell staining vs tumour-cell staining
There is generally higher agreement between observers when assessing TPS than when assessing ICPS

41. Blueprint Project: Step 1 Study Results
Analytical comparison of tumor and immune-cell–membrane staining for each of the 39 NSCLC cases, using all four PD-L1 assays 60 70 80 90
100
% Tumour Cell Staining 10 20 30 40 50
Cases 1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39
22C3
28-8
SP142
SP263
Tumour Cells
% Immune Cell Staining
Cases 1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 60 70 80 90
100 10 20 30 40 50
22C3
28-8
SP142
SP263
Immune Cells
Data points represent the mean score from three pathologists, for each assay, with each NSCLC case
Identical PD-L1 expression reported is represented by superimposed points
No clinically relevant cutoff is applied

42. Blueprint Project: Clinical Diagnostic Comparison
36.9% of the cases studied showed discrepant results for PD-L1 expression between the assays
There is the potential for different diagnostic results according to the key clinical cutoffs if assays and algorithms are mismatched
The assays, the cutoffs, and the therapeutics may not be interchangeable
The results of this preliminary study should not alter current guidelines as indicated for each therapeutic-diagnostic validated combination pair
Further studies are required 19 6 5 1
SP142 TC1IC1
(30)
28-8 1%
(26)
22C3 1%
SP263 25%
(20)
All four assays are in agreement for 19 cases
Assay
Overall agreement with each selected cutoff
1% TPS
TC1/IC1
25% TPS
22C3
38/38 (100%)
31/38 (81.6%)
30/38 (78.9%)
28-8
29/38 (76.3%)
SP142
20/38 (52.6%)
15/38 (39.5%)
SP263
32/38 (84.2%)
23/38 (60.5%)
Agreement rates between assays and cutoffs are shown

43. Courtesy of Professor Fred Hirsch
The Blue Print Project
Courtesy of Professor Fred Hirsch

44. Practical Considerations for PD-L1 Testing
IVDs vs LDTs
Formal HTA Assessment of PD-L1 Dx Test Is Not Required in Most Markets
Class III In Vitro Diagnostics1,2
Developed in the form of a kit for general use
FDA approval of kits required
CE marking required
Highly regulated PMA process before being marketed
Clinical safety or performance evaluation required
Laboratory-Developed Tests1,2
Created by local laboratories, at which use is restricted
Created by local laboratories
No requirement for premarket review, plans in place for future
No requirement for premarket review, manufacturer self-declares
Clinical validation is not required for their use
Clinical safety or performance evaluation required
CE = Conformité Européene (European Conformity); Dx = diagnostic; HER2 = human epidermal growth factor receptor 2; HTA = health technology assessment; IVDs = in vitro diagnostics; LDTs = laboratory-developed tests; PMA = premarket approval.
1. In Vitro Companion Diagnostic Devices. FDA Guidance. August 6, Accessed September Understanding Europe’s New Medical Devices Regulation (MDR). Key changes contained in the proposed MDR and their impact on manufacturers. EMERGO white paper: July 2016, anticipated publishing date early 2017.

45. Assays and LDTs: Mix and Match?
Drug
Anti–PD-L1 Assay
Definition of Positivity
Outcome
Risk to Patient?
By choice
Assay validated for drug in trial
Definition validated in trial for drug
Predictable based upon trial data
Known
Any trial-validated assay
Definition validated in trial for drug of choice
Uncertain
Not known
Definition validated with the assay
Very uncertain
LDT using any clone
Unknown
Very, very uncertain
Courtesy of Professor Keith Kerr

46. The “Consensus” we may end up with…
Cancer Patients (to be treated by Anti-PD1 / Anti-PDL1)
Reflex Testing?
PDL1 IHC accompanied with / complimentary to Drug A
PDL1 IHC for Drug B
PDL1 IHC for Drug C + + +
Drug B
Drug A
Drug C

47. Goals:
Compare analytical performance of 4 assays (22C3, 28-8, SP142, SP263) using the staining protocols used in respective clinical trials
Compare the treatment-determining scoring algorithm developed for each assay and used in clinical trials
Journal of Thoracic Oncology Vol. 12 No. 2:

48. Blueprint phase 2 - Goals
Validation of Blueprint phase 1 results using different types of clinical samples (resection, small biopsy, cytology cell block)
Comparability and heterogeneity of PD-L1 assay results in surgical tumor resection, core needle and FNA samples prepared from same tumor.
Inter-observers concordance among larger panel of pulmonary pathologists
Concordance of PD-L1 scoring using standard light microscopy vs. digital images accessed by web-based system.

49. Blueprint Phase 2 (2017) FDA EMA BMS Steering Committee PFE/EMD Serono
Astra-Zeneca
BMS
Genentech
Merck
Funders
Technical facilitator
Dako
Ventana
IASLC
Pathology expertise
Execution Team
HistoGeneX
Technical expertise
Core Team
Regulatory/public advocates
Neutral observers
FDA
EMA

50. There is literature comparing different PD-L1 IHC assays but……
Mostly of limited value in the testing debate
Based upon LDTs
Tell us nothing about the comparability of trial-validated assays
May not be clinically practical
BUT (and inevitably)
Many demonstrate differences in outcome on the same cases

51. IASLC Atlas of PD-L1 IHC Testing

53. The Need of Liquid Biopsies in Oncology
Diagnosis of Cancer
Prognosis Prediction
Therapy Response Prediction
Risk Assessment for Relapse
Monitoring Tumor Burden
Treatment Selection

54. Liquid Biopsies Capture Molecular Heterogeneity of Tumor

55. PDAC CTC Cluster Count Criteria
Sub-classification of PDAC CTC Cluster
Sub-classification #1
CTC cluster contain = 1 CTC
Sub-classification #2
CTC cluster contain = 2 CTC
Sub-classification #3
CTC cluster contain = 3 CTC
CTC cluster contain >= 5 cells
CTC Cluster Criteria
Contain one CTC with WBC
Size > 15um
Contact nuclear stain
Contain 2 CTC in any shape with x WBCs
Size >20um
Contain 3 CTC in any shape with x WBCs
Size >25um
CTC cluster more than 5 cells contain at least one CTC
Represented Figures
Red: PanCK
Green: CD45
Blue: DAPI