Case 1:20-cv-01580-LPS Document 1-14 Filed 11/23/20 Page 1 of 36 PageID #: 490
`Case 1:20-cv-01580-LPS Document 1-14 Filed 11/23/20 Page 1 of 36 PageID #: 490
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`EXHIBIT 14
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`EXHIBIT 14
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`

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`Case 1:20-cv-01580-LPS Document 1-14 Filed 11/23/20 Page 2 of 36 PageID #: 491
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`FoundationOne® Liquid CDx
`Technical Information
`
`Foundation Medicine, Inc.
`150 Second Street, Cambridge, MA 02141
`Phone: 617.418.2200
`
`Intended Use
`FoundationOne Liquid CDx is a qualitative next generation sequencing based in vitro diagnostic test that uses
`targeted high throughput hybridization-based capture technology to detect and report substitutions, insertions and
`deletions (indels) in 311 genes, including rearrangements and copy number losses only in BRCA1 and BRCA2.
`FoundationOne Liquid CDx utilizes circulating cell-free DNA (cfDNA) isolated from plasma derived from anti-
`coagulated peripheral whole blood of cancer patients collected in FoundationOne Liquid CDx cfDNA blood
`collection tubes included in the FoundationOne Liquid CDx Blood Sample Collection Kit. The test is intended to
`be used as a companion diagnostic to identify patients who may benefit from treatment with the targeted therapies
`listed in Table 1 in accordance with the approved therapeutic product labeling. Additionally, FoundationOne Liquid
`CDx is intended to provide tumor mutation profiling for substitutions and indels to be used by qualified health care
`professionals in accordance with professional guidelines in oncology for patients with solid malignant neoplasms.
`
`Table 1: Companion diagnostic indications
`Tumor Type
`Biomarker(s) Detected
`
`Non-small cell lung
`cancer (NSCLC)
`
`EGFR Exon 19 deletions and
`EGFR Exon 21 L858R substitution
`
`Prostate cancer
`
`BRCA1, BRCA2 alterations
`
`Therapy
`IRESSA® (gefitinib)
`TAGRISSO® (osimertinib)
`TARCEVA® (erlotinib)
`RUBRACA® (rucaparib)
`
` A
`
` negative result from a plasma specimen does not mean that the patient’s tumor is negative for genomic findings.
`Patients who are negative for the mutations listed in Table 1 should be reflexed to routine biopsy and their tumor
`mutation status confirmed using an FDA-approved tumor tissue test, if feasible.
`
`Genomic findings other than those listed in Table 1 are not prescriptive or conclusive for labeled use of any
`specific therapeutic product.
`
`
`FoundationOne Liquid CDx is a single-site assay performed at Foundation Medicine, Inc. in Cambridge, MA.
`
`Contraindication
`There are no known contraindications.
`
`Warnings and Precautions
`• Alterations reported may include somatic (not inherited) or germline (inherited) alterations; however, the
`test does not distinguish between germline and somatic alterations. If a reported alteration is suspected
`to be germline, confirmatory testing should be considered in the appropriate clinical context.
`
`
`
`
`
`
`
`• The test is not intended to replace germline testing or to provide information about cancer
`predisposition.
`
`• Patients for whom no companion diagnostic alterations are detected should be considered for
`confirmation with an FDA-approve tumor tissue test, if possible.
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`Page 1 of 35
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`Case 1:20-cv-01580-LPS Document 1-14 Filed 11/23/20 Page 3 of 36 PageID #: 492
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`Limitations
`• For in vitro diagnostic use.
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`• For prescription use only. This test must be ordered by a qualified medical professional in accordance
`with clinical laboratory regulations.
`
`• Genomic findings other than those listed in Table 1 of the intended use are not prescriptive or conclusive
`for labeled use of any specific therapeutic product.
`
`• A negative result does not rule out the presence of a mutation in the patient’s tumor.
`
`• Decisions on patient care and treatment must be based on the independent medical judgment of the
`treating physician, taking into consideration all applicable information concerning the patient’s condition,
`such as patient and family history, physical examinations, information from other diagnostic tests, and
`patient preferences, in accordance with the standard of care in a given communities.
`
`• The test is intended to be performed on specific serial number-controlled instruments by Foundation
`Medicine, Inc.
`
`• Genomic findings from circulating cell-free DNA (cfDNA) may originate from circulating tumor DNA
`fragments, germline alterations, or non-tumor somatic alterations, such as clonal hematopoiesis of
`indeterminate potential (CHIP). Genes with alterations that may be derived from CHIP include, but are not
`limited to: ASXL1, ATM, CBL, CHEK2, DNMT3A, JAK2, KMT2D (MLL2), MPL, MYD88, SF3B1, TET2,
`TP53, and U2AF1.
`
`• The false positive rate of this test was evaluated in healthy donors. The detection rate for unique short
`variants in apparently healthy patients is 0.82%. Across 30,622 short variants, 58 variants had a detection
`rate of greater than 5%.
`
`• The analytical accuracy for the FoundationOne Liquid CDx assay has not been demonstrated in all genes.
`
`• The precision of FoundationOne Liquid CDx was only confirmed for select variants at the limit of detection.
`
`• The FoundationOne Liquid CDx assay does not detect heterozygous deletions.
`
`• A complete assessment of the impact of cfDNA blood collection tube lot-to-lot variability on the
`performance of the test has not been evaluated.
`
`• The test is not intended to provide information on cancer predisposition.
`
`• BRCA1/BRCA2 homozygous deletions and rearrangements were not adequately represented in all
`analytical studies.
`
`• Performance has not been validated for cfDNA input below the specified minimum input.
`
`
`Test Principle
`The FoundationOne Liquid CDx assay is performed exclusively as a laboratory service using circulating cell-
`free DNA (cfDNA) isolated from plasma derived from anti-coagulated peripheral whole blood from patients with
`solid malignant neoplasms. The assay employs a single DNA extraction method to obtain cfDNA from plasma
`from whole blood. Extracted cfDNA undergoes whole-genome shotgun library construction and hybridization-
`based capture of 324 cancer-related genes. All coding exons of 309 genes are targeted; select intronic or non-
`coding regions are targeted in BRCA1 and BRCA2 (refer to Table 2 for the complete list of genes reported by
`FoundationOne Liquid CDx). Hybrid-capture selected libraries are sequenced with deep coverage using the
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`Case 1:20-cv-01580-LPS Document 1-14 Filed 11/23/20 Page 4 of 36 PageID #: 493
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`NovaSeq® 6000 platform. Sequence data are processed using a custom analysis pipeline designed to detect
`genomic alterations, including base substitutions and indels in 311 genes, and copy number variants and
`genomic rearrangements in BRCA1 and BRCA2. A subset of targeted regions in 75 genes is baited for
`increased sensitivity.
`
`Table 2: As part of its FDA-approved intended use, the FoundationOne Liquid CDx assay interrogates 311
`genes, including 309 genes with complete exonic (coding) coverage and 2 genes with only select non-
`coding coverage (indicated with an *).
`
`Select genes and select exons (indicated in bold) are captured with increased sensitivity.
`
`ABL1 [Exons
`4-9]
`
`CALR
`
`CYP17A1
`
`FGFR4
`
`KDM6A
`
`MYCL
`(MYCL1)
`
`POLD1
`
`SMAD4
`
`ACVR1B
`
`CARD11
`
`DAXX
`
`FH
`
`KDR
`
`MYCN
`
`POLE
`
`SMARCA4
`
`AKT1 [Exon
`3]
`
`CASP8
`
`DDR1
`
`AKT2
`
`CBFB
`
`DDR2 [Exons
`5,17,18]
`
`FLCN
`
`FLT1
`
`KEAP1
`
`MYD88
`[Exon 4]
`
`PPARG
`
`SMARCB1
`
`KEL
`
`NBN
`
`PPP2R1A
`
`SMO
`
`AKT3
`
`CBL
`
`DIS3
`
`FLT3 [Exons
`14,15,20]
`
`KIT [Exons
`8,9,11,12,13
`,17]
`
`CCND1
`
`DNMT3A
`
`FOXL2
`
`CCND2
`
`DOT1L
`
`FUBP1
`
`CCND3
`
`EED
`
`GABRA6
`
`KLHL6
`
`KMT2A
`(MLL)
`KMT2D
`(MLL2)
`
`ALK [Exons
`20-29]
`
`ALOX12B
`
`
`AMER1
`(FAM123B)
`
`APC
`
`AR
`
`NF1
`
`NF2
`
`PPP2R2A
`
`SNCAIP
`
`PRDM1
`
`SOCS1
`
`NFE2L2
`
`PRKAR1A
`
`SOX2
`
`NFKBIA
`
`PRKCI
`
`SOX9
`
`SPEN
`
`SPOP
`
`CCNE1
`
`EGFR
`
`GATA3
`
`KRAS
`
`NKX2-1
`
`PTCH1
`
`CD22
`
`EP300
`
`GATA4
`
`LTK
`
`NOTCH1
`
`PTEN
`
`EPHA3
`
`GATA6
`
`LYN
`
`NOTCH2
`
`PTPN11
`
`SRC
`
`ARAF
`[Exons 4,5,7,
`11,13,15,16]
`
`CD274
`(PD-L1)
`
`ARFRP1
`
`CD70
`
`EPHB1
`
`GNA11
`[Exons 4,5]
`
`MAF
`
`NOTCH3
`
`PTPRO
`
`STAG2
`
`ARID1A
`
`CD79A
`
`EPHB4
`
`GNA13
`
`ASXL1
`
`CD79B
`
`ERBB2
`
`GNAQ
`[Exons 4,5]
`
`MAP2K1
`(MEK1)
`[Exons 2,3]
`MAP2K2
`(MEK2)
`[Exons 2-
`4,6,7]
`
`NPM1
`[Exons 4-
`6,8,10]
`
`QKI
`
`STAT3
`
`NRAS [Exons
`2,3]
`
`RAC1
`
`STK11
`
`ATM
`
`CDC73
`
`ERBB3
`[Exons 3,6,7,
`8,10,12,20,2
`1,23,24,25]
`
`GNAS
`[Exons 1,8]
`
`MAP2K4
`
`NSD3
`(WHSC1L1)
`
`RAD21
`
`SUFU
`
`ATR
`
`CDH1
`
`ERBB4
`
`GRM3
`
`MAP3K1
`
`ATRX
`
`CDK12
`
`ERCC4
`
`GSK3B
`
`MAP3K13
`
`AURKA
`
`CDK4
`
`ERG
`
`H3F3A
`
`MAPK1
`
`AURKB
`
`CDK6
`
`ERRFI1
`
`HDAC1
`
`MCL1
`
`AXIN1
`
`CDK8
`
`ESR1
`[Exons 4-8]
`
`HGF
`
`MDM2
`
`NT5C2
`
`NTRK1
`[Exons
`14,15]
`
`NTRK2
`
`NTRK3
`[Exons
`16,17]
`
`P2RY8
`
`RAD51
`
`SYK
`
`RAD51B
`
`TBX3
`
`RAD51C
`
`TEK
`
`RAD51D
`
`RAD52
`
`TERC*
`{ncRNA}
`
`TERT*
`{Promoter}
`
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`AXL
`
`CDKN1A
`
`EZH2 [Exons
`4,16,17,18]
`
`HNF1A
`
`MDM4
`
`PALB2
`
`RAD54L
`
`TET2
`
`BAP1
`
`CDKN1B
`
`FAM46C
`
`HRAS [Exons
`2,3]
`
`MED12
`
`PARK2
`
`RAF1 [Exons
`3,4,6,7,10,1
`4,15,17]
`
`TGFBR2
`
`BARD1
`
`CDKN2A
`
`FANCA
`
`HSD3B1
`
`MEF2B
`
`PARP1
`
`RARA
`
`TIPARP
`
`BCL2
`
`CDKN2B
`
`FANCC
`
`ID3
`
`MEN1
`
`PARP2
`
`RB1
`
`TNFAIP3
`
`MERTK
`
`PARP3
`
`RBM10
`
`TNFRSF14
`
`BCL2L1
`
`CDKN2C
`
`FANCG
`
`BCL2L2
`
`CEBPA
`
`FANCL
`
`IDH1 [Exon
`4]
`IDH2 [Exon
`4]
`
`CHEK1
`
`FAS
`
`IGF1R
`
`BCL6
`
`BCOR
`
`MET
`
`MITF
`
`CHEK2
`
`FBXW7
`
`IKBKE
`
`MKNK1
`
`BCORL1
`
`CIC
`
`FGF10
`
`IKZF1
`
`MLH1
`
`BRAF [Exons
`11-18]
`
`CREBBP
`
`FGF12
`
`INPP4B
`
`MPL [Exon
`10]
`
`PAX5
`
`REL
`
`TP53
`
`TSC1
`
`RET [Exons
`11,13-16]
`
`RICTOR
`
`TSC2
`
`RNF43
`
`TYRO3
`
`ROS1 [Exons
`31,36-38,40]
`
`U2AF1
`
`PBRM1
`
`PDCD1
`(PD-1)
`PDCD1LG2
`(PD-L2)
`PDGFRA
`[Exons
`12,18]
`
`FGF14
`
`IRF2
`
`MRE11A
`
`RPTOR
`
`VEGFA
`
`BRCA1
`{Introns 2, 7,
`8, 12, 16, 19,
`20}
`BRCA2
`{Intron 2}
`
`BRD4
`
`BRIP1
`
`CRKL
`
`PDGFRB
`[Exons 12-
`21,23]
`
`CSF1R
`
`FGF19
`
`CSF3R
`
`FGF23
`
`CTCF
`
`FGF3
`
`IRF4
`
`IRS2
`
`JAK1
`
`MSH2
`
`PDK1
`
`SDHA
`
`VHL
`
`MSH3
`
`PIK3C2B
`
`SDHB
`
`WHSC1
`
`MSH6
`
`PIK3C2G
`
`SDHC
`
`WT1
`
`BTG1
`
`CTNNA1
`
`FGF4
`
`JAK2 [Exons
`14]
`
`MST1R
`
`PIK3CA
`[Exons
`2,3,5-
`8,10,14,19,2
`1] (Coding
`Exons 1, 2,
`4-7, 9, 13,
`18, 20)
`
`SDHD
`
`XPO1
`
`BTG2
`
`CTNNB1
`[Exon 3]
`
`FGF6
`
`JAK3 [Exons
`5,11,12,13,1
`5,16]
`
`MTAP
`
`PIK3CB
`
`SETD2
`
`XRCC2
`
`CUL3
`
`FGFR1
`
`JUN
`
`MTOR
`[Exons
`19,30,39,40,
`43-
`45,47,48,53,
`56]
`
`PIK3R1
`
`SF3B1
`
`ZNF217
`
`CUL4A
`
`FGFR2
`
`KDM5A
`
`MUTYH
`
`PIM1
`
`SGK1
`
`ZNF703
`
`BTK [Exons
`2,15]
`
`C11orf30
`(EMSY)
`
`C17orf39
`(GID4)
`
`CXCR4
`
`
`
`FGFR3
`[Exons 7, 9
`(alternative
`designation
`exon 10),14,
`18]
`
`KDM5C
`
`MYC
`
`PMS2
`
`SMAD2
`
`
`
`Table 3 and Table 4 describe the criteria for classifying BRCA1 or BRCA2 alterations known to be deleterious to
`BRCA protein function rendering the sample BRCA+.
`
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`Table 3: Classification Criteria for Deleterious Tumor BRCA Variants
`Qualification
`Sequence
`Methodology
`Criteria
`Classification
`Protein truncating
`A BRCA1/2
`mutations
`alteration that
`includes any of
`the sequence
`classifications
`
`Sequence analysis identifies
`premature stop codons
`anywhere in the gene coding
`region, except: 3’ of and
`including BRCA2 K3326*
`Sequence analysis identifies
`variant splice sequences at
`intron/exon junctions -/+ 2bp
`of exon starts/ends
`Sequence analysis identifies
`deletions in both gene alleles
`of ≥ 1 exon in size
`Sequence analysis identifies
`protein truncating
`rearrangements
`Curated list (Table 4)
`
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`Splice site mutations
`
`Homozygous deletions
`
`Large protein truncating
`rearrangements
`
`Deleterious missense
`mutations
`
`Table 4: Deleterious BRCA Missense Alterations
`BRCA1 Alterations
`BRCA2 Alterations
`(Protein Change)
`(Protein Change)
`M1V
`C61G
`D1692H G1788V M1V
`R2659T
`
`M1T
`
`M1R
`
`M1I
`
`C61Y
`
`C64R
`
`C64G
`
`D1692Y P1812A M1T
`
`R1699W A1823T M1R
`
`R1699Q V1833M M1I
`
`R2659K
`
`E2663V
`
`S2670L
`
`M18T C64Y
`
`G1706R W1837R D23N
`
`I2675V
`
`L22S C64W
`
`G1706E V1838E D23Y
`
`T2722K
`
`I26N
`
`R71G
`
`A1708E
`
`T37K R71K
`
`S1715R
`
`C39R R71T
`
`S1722F
`
`C39G R71M
`
`V1736A
`
`
`
`
`
`
`
`
`
`
`
`S142N
`
`T2722R
`
`S142I
`
`D2723H
`
`V159M
`
`D2723G
`
`V211I
`
`G2724W
`
`C39Y S770L
`
`G1738R
`
`C39W R1495T G1738E
`
`H41R R1495M K1759N
`
`C44S R1495K L1764P
`
`C44Y E1559K
`
`I1766N
`
`C44F E1559Q
`
`I1766S
`
`C47S T1685A G1770V
`
`C47Y T1685I M1775K
`
`C47F D1692N M1775R
`
`C61S M1689R C1787S
`
`
`
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`
`
`V211L
`
`G2748D
`
`Y600C
`
`A2911E
`
`K1530N E3002K
`
`R2336P R3052W
`
`R2336L D3095G
`
`R2336H D3095E
`
`T2412I
`
`N3124I
`
`R2602T N3187K
`
`W2626C
`
`I2627F
`
`
`
`
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`The output of the test includes:
`
`
`Category 1: Companion Diagnostic (CDx) claims noted in Table 1 of the Intended Use
`
`Category 2: cfDNA Biomarkers with Strong Evidence of Clinical Significance in cfDNA
`
`Category 3: Biomarkers with Evidence of Clinical Significance in tissue supported by:
`3A: strong analytical validation using cfDNA
`3B: analytical validation using cfDNA
`
`
`Category 4: Other Biomarkers with Potential Clinical Significance
`
`
`FoundationOne Liquid CDx cfDNA Blood Specimen Collection Kit Contents
`
`Test Kit Contents
`The test includes a sample shipping kit, which is sent to ordering laboratories. The shipping kit contains the
`following components:
`• Specimen preparation and shipping instructions
`• Two FoundationOne Liquid CDx cfDNA blood collection tubes (8.5 mL nominal fill volume per tube)
`• Return shipping label
`
`
`All other reagents, materials and equipment needed to perform the assay are used exclusively in the Foundation
`Medicine laboratory. The FoundationOne Liquid CDx assay is intended to be performed with serial number-
`controlled instruments.
`
`FoundationOne Liquid CDx Sample Collection and Test Ordering
`To order FoundationOne Liquid CDx, the test order form in the test kit must be fully completed and signed by the
`ordering physician or other authorized medical professional. Please refer to Specimen Preparation Instructions
`and Shipping Instructions included in the test kit.
`
`For more detailed information, including Performance Characteristics, please find the FDA Summary of Safety
`and Effectiveness Data at: [Placeholder]
`
`Instruments
`The FoundationOne Liquid CDx device is intended to be performed with the following instruments, as identified
`by specific serial numbers:
`Illumina NovaSeq 6000
`•
`• Beckman Biomek NXP Span-8 Liquid Handler
`• Thermo Scientific Kingfisher Flex DW 96
`• Bravo Benchbot
`• Hamilton STARTlet-STAR Liquid Handling Workstation
`
`1.
`
`
`2. Performance Characteristics
`Performance characteristics were established using contrived and clinical circulating cfDNA derived from blood
`specimens extracted from a wide range of tumor types. Table 5 below provides a summary of the number of
`tumor types and variants included in each study. As summarized in this table, each study included a broad range
`of representative alteration types (substitutions, insertion-deletions, copy number alterations, rearrangements) in
`various genomic contexts across a number of genes. The validation studies included >7,000 sample replicates,
`>31,000 unique variants [includes variants classified as variants of unknown significance (VUS) and/or benign],
`>30 tumor types, representing all 324 genes targeted by the assay.
`
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`Table 5. Representation of tumor types and variants across validation studies
`# of Unique
`
`Study Title
`
`Cancer Types
`Represented
`
`# Unique
`Samples
`
`# of
`Sample
`Replicates
`
`Targeted
`Genes
`
`Subs
`
`Indels
`
`Rearrang.
`
`Copy
`Number
`Losses
`
`Contrived Sample
`Functional
`Characterization
`(CSFC) Study
`FoundationOne
`Liquid CDx to
`Validated NGS
`Tumor Tissue Test
`Concordance:
`BRCA1 and BRCA2
`Variants
`Orthogonal
`Concordance
`
`LoD Estimation
`
`Breast cancer
`Colorectal cancer
`Lung cancer
`Contrived samples
`
`Prostate cancer
`Ovarian cancer
`
`23 cancer types
`Contrived samples
`Prostate
`Contrived samples
`Healthy Donors
`
`Contrived samples
`
`25 cancer types
`Contrived samples
`Contrived samples
`
`13
`
`1843
`
`228
`
`563
`
`81
`
`11
`
`1
`
`279
`
`N/A
`
`2
`
`100
`
`87
`
`9
`
`2
`
`278
`
`10
`
`28
`
`9
`
`3546
`
`8
`
`N/A
`
`877
`
`79
`
`336
`
`N/A
`
`142
`
`11
`
`32
`
`64
`
`541
`
`12
`
`286
`
`322
`
`1490
`
`247
`
`26134
`
`4482
`
`911
`
`18
`
`16
`
`11
`
`11
`
`324
`
`279
`
`N/A
`
`1090
`
`N/A
`
`215
`
`11
`
`N/A
`
`32
`
`11
`
`0
`
`3
`
`42
`
`2
`
`N/A
`
`2
`
`1
`
`LoB
`Potentially
`Interfering
`Substances
`Hybrid Capture Bait
`Specificity
`Reagent Stability
`Reagent
`Interchangeability
`
`Precision study 1
`
`Precision study 2
`
`DNA Extraction
`
`Whole Blood
`Sample Stability
`
`Inverted Tube
`Whole Blood
`Sample Stability
`
`Cross
`Contamination
`Guard Banding
`
`Contrived samples
`
`8
`
`192
`
`20
`
`15
`
`Breast cancer
`Colon cancer
`Lung cancer
`Ovarian cancer
`Prostate cancer
`Skin cancer
`Contrived samples
`Lung cancer
`Prostate cancer
`Stomach cancer
`Colorectal cancer
`Bile duct cancer
`Breast cancer
`Colorectal cancer
`Prostate cancer
`Breast cancer
`Lung cancer
`Skin cancer
`Lung cancer
`Colorectal cancer
`Prostate cancer
`Breast cancer
`Lung cancer
`Colorectal cancer
`Breast cancer
`Ovarian cancer
`Prostate cancer
`
`Contrived samples
`
`Contrived samples
`
`47
`
`1121
`
`280
`
`900
`
`229
`
`63
`
`5
`
`10
`
`230
`
`6
`
`6
`
`4
`
`0
`
`0
`
`6
`
`72
`
`161
`
`265
`
`53
`
`11
`
`22
`
`66
`
`75
`
`15
`
`130
`
`260
`
`237
`
`594
`
`91
`
`5
`
`10
`
`376
`
`375
`
`39
`
`20
`
`9
`
`17
`
`5
`
`12
`
`2
`
`1
`
`5
`
`4
`
`12
`
`0
`
`0
`
`0
`
`1
`
`1
`
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`Study Title
`
`Cancer Types
`Represented
`
`# Unique
`Samples
`
`# of
`Sample
`Replicates
`
`Targeted
`Genes
`
`# of Unique
`
`Subs
`
`Indels
`
`Rearrang.
`
`Copy
`Number
`Losses
`
`Clinical validation
`for detection of
`EGFR exon 19
`deletions and
`L858R alterations:
`non-inferiority study
`Clinical validation
`study for detection
`of deleterious
`alterations in
`BRCA1 and BRCA2
`in prostate cancer
`
`Blood Collection
`Tube Equivalence
`
`Automation Line
`Equivalence
`
`Variant Report
`Curation
`
`Lung cancer
`
`177
`
`N/A
`
`1
`
`5
`
`7
`
`N/A
`
`N/A
`
`Prostate cancer
`
`199
`
`N/A
`
`2
`
`44
`
`55
`
`8
`
`1
`
`Ovarian cancer
`Breast cancer
`Colorectal cancer
`Prostate cancer
`Lung cancer
`Skin cancer
`Stomach cancer
`
`60
`
`192
`
`116
`
`135
`
`39
`
`13
`
`Contrived samples
`
`8
`
`187
`
`303
`
`1926
`
`337
`
`63
`
`Breast cancer
`Colorectal cancer
`Lung cancer
`Prostate cancer
`Skin cancer
`
`19
`
`57
`
`183
`
`300
`
`104
`
`15
`
`0
`
`4
`
`2
`
`20 cancer types
`
`19868
`
`N/A
`
`25 cancer types
`Contrived samples
`
`7637
`
`N/A
`
`Pan-tumor
`performance
`(includes historical
`analysis)
`Molecular Index
`Barcode
`Performance
`FoundationOne
`Liquid LDT to
`FoundationOne
`Liquid CDx
`Concordance
`* Variants detected include variants classified as VUS and benign.
`
`25 cancer types
`
`927
`
`N/A
`
`N/A
`
`N/A
`
`N/A
`
`N/A
`
`N/A
`
`324
`
`N/A
`
`N/A
`
`N/A
`
`N/A
`
`73
`
`1815
`
`376
`
`109
`
`N/A
`
`
`2.1 Concordance – Comparison to an Orthogonal cfDNA NGS Method #1
`The detection of short variants and rearrangements by the FoundationOne Liquid CDx assay was compared to
`that of an externally validated NGS assay in 74 genes common to both assays across 278 samples that
`represented an array of tumor types (>50 unique disease ontologies across 23 cancer types, Table 9). The cancer
`types (# samples) included lung [NSCLC (75) and other (3)]; breast (54); prostate (32); colorectal [colon (27) and
`rectal (6)]; liver (11); ovarian (6); pancreas (9); gastrointestinal (7); bile duct (2); esophageal (5); skin (6); cervical
`(1); anal (1); bladder (1); gallbladder (1); salivary gland (2); thymus (1); thyroid (3); uterine (2); fallopian tube (1);
`head and neck (1); soft tissue (1); and unknown primary (19). The study included samples selected from clinical
`FoundationOne Liquid testing (n=268) and contrived samples consisting of fragmented gDNA diluted in clinical
`cfDNA to represent rare alterations (n=10).
`
`Using the externally validated NGS assay as the comparator, the analysis demonstrated a short variant PPA of
`96.2% with a 95% two-sided CI of [94.8%-97.4%]. The short variant NPA was >99.9% with a 95% two-sided CI
`of [99.9%-100.0%]. The respective PPA of base substitutions and indels with a 95% two-sided CI was 96.1%
`[94.6%-97.3%] and 100.0% [85.2%-100.0%]. The respective NPA and 95% two-sided CI of base substitutions
`and indels was >99.9% [99.9%-100.0%] and 100.0% [99.89%-100.0%] (Table 6).
`
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`Table 6. Concordance of short variants called in FoundationOne Liquid CDx and the comparator assay
`(n= 902 positive variants, n= 152,832 negative variants* by the comparator assay)
`
`PPA
`[95% CI]
`
`NPA
`[95% CI]
`
`OPA
`[95% CI]
`
`Comparator(-)
`Liquid CDx(-)
`FoundationOne
`
`Comparator(-)
`Liquid CDx(+)
`FoundationOne
`
`Comparator(+)
`Liquid CDx(-)
`FoundationOne
`
`Comparator(+)
`Liquid CDx(+)
`FoundationOne
`
`Variant Type
`
`All Short
`Variants
`Base
`Substitutions
`
`868
`
`845
`
`Indels
`
`23
`
`34
`
`34
`
`0
`
`8
`
`8
`
`0
`
`152824
`
`149511
`
`3361
`
`96.2%
`[94.8%-97.4%]
`96.2%
`[94.6%-97.3%]
`100.0%
`[85.2%- 100.0%]
`* Variants detected include variants classified as VUS and benign.
`
`For the concordance of rearrangement detection between FoundationOne Liquid CDx and the comparator assay,
`the observed rearrangement PPA was 100.0%, with a 95% two-sided CI of [59.0%-100.0%]. The NPA was 99.8%,
`with a 95% two-sided CI [99.5%-100.0%] (Table 7).
`
`Table 7. Concordance of rearrangements called in FoundationOne Liquid CDx and the comparator assay
`(n= 7 positive, n=1685 negative* as determined by the comparator assay)
`
`>99.9%
`[99.9%-100.0%]
`>99.9%
`[99.9%-100.0%]
`100.0%
`[99.9%- 100.0%]
`
`>99.9%
`[99.9%-100.0%]
`>99.9%
`[99.9%-100.0%]
`100.0%
`[99.9%- 100.0%]
`
`
`
`Comparator (+)
`
`Comparator (-)
`
`FoundationOne Liquid CDx (+)
`
`FoundationOne Liquid CDx (-)
`
`Total
`
`
`
`7
`
`0
`
`7
`
`PPA:
`
`3
`
`1682
`
`1685
`
`NPA:
`
`Total
`
`10
`
`1682
`
`1692
`
`OPA:
`
`100.0%
`[59.0% - 100.0%]
`
`99.8%
`[99.5% - 100.0%]
`
`99.8%
`[99.5% - 100.0%]
`
`* Variants detected include variants classified as VUS and benign.
`
`Assessment of a subset of highly-actionable alterations were compared between the two assays. The analysis
`resulted in a PPA of 100% across all eligible highly-actionable alterations called in the comparator assay (Table
`8).
`
`Table 8. Concordance of highly actionable alterations called between FoundationOne Liquid CDx and the
`comparator assay (n = 73)
`
`Targeted Alteration
`
`BRCA1 short variants
`
`BRCA2 short variants
`
`n
`
`1
`
`2
`
`PPA [95% CI]
`
`NPA [95% CI]
`
`100% [2.5%-100.0%]
`
`100% [98.7%-100.0%]
`
`100% [15.8%-100.0%] 100% [99.3%-100.0%]
`
`EGFR exon 19 deletions
`
`11
`
`100% [71.5%-100.0%] 100% [99.7%-100.0%]
`
`EGFR L858R
`
`10
`
`100% [69.2%-100.0%] 100% [98.7%-100.0%]
`
`
`
`These data demonstrate that the FoundationOne Liquid CDx assay and an externally-validated NGS assay are
`highly concordant across the 74 genes common between the two panels.
`
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`2.2 Concordance – FoundationeOne Liquid CDx to validated NGS tumor tissue assay (BRCA1 and BRCA2
`alterations)
`Samples from a total of 279 prostate and ovarian cancer patients were tested and the concordance evaluated
`between FoundationOne Liquid CDx and the validated NGS tumor tissue assay for the detection of deleterious
`alterations in BRCA1 or BRCA2. As summarized below, a PPA of 88.03% and an NPA of 95.68% were observed
`on a sample level (Table 9). As summarized in Table 10, an overall PPA of 87.28% and an NPA of 99.83% were
`observed at the variant level. Some discordance is expected based on biological differences and sampling times
`between tumor tissue and plasma samples. Considering the impact of biological differences between analytes,
`these data demonstrate a high concordance between FoundationOne Liquid CDx and FoundationOne for the
`detection of deleterious alterations in BRCA1 or BRCA2.
`
`Table 9. Concordance (by sample) of FoundationOne Liquid CDx and validated NGS tumor tissue assay
`in prostate and ovarian cancer patients for the detection of alterations in BRCA1 or BRCA2
`
`NGS Tumor Tissue Assay
`
`Positive
`Negative
`
`FoundationOne Liquid CDx
`
`Positive
`
`Negative
`
`103
`
`14
`
`7
`
`155
`
`
`
`
`
`PPA: 88.03%
`[80.91%-92.74%]
`
`NPA: 95.68%
`[91.35%-97.89%]
`
`Table 10. Concordance (by variant) of FoundationOne Liquid CDx and validated NGS tumor tissue
`assay in prostate and ovarian cancer patients for the detection of alterations in BRCA1 or BRCA2
`F1LCDx+
`F1L CDx-
`F1L CDx+
`F1L CDx-/
`PPA (%)
`NPA (%)
`/Tissue+
`/Tissue+
`/Tissue-
`Tissue-
`CI1
`CI1
`92.77
`99.86
`(85.11, 96.64)
`(99.79, 99.90)
`95.59
`99.81
`(87.81, 98.49)
`(99.73, 99.87)
`57.14
`99.64
`(25.05, 84.18)
`(99.26, 99.83)
`33.33
`99.62
`(15.18, 58.29)
`(97.89, 99.93)
`87.28
`99.83
`(81.50, 91.45)
`(99.78, 99.86)
`
`6
`
`3
`
`3
`
`10
`
`22
`
`29
`
`31
`
`7
`
`1
`
`20255
`
`16362
`
`1939
`
`263
`
`68
`
`38819
`
`
`
`Substitutions
`
`Indels
`
`Rearrangements
`
`Copy number
`loss
`
`77
`
`65
`
`4
`
`5
`
`Total
`
`151
`
`
`2.3 Limit of Detection (Analytical Sensitivity)
`The LoD for each variant type was established by processing a total of 1,069 sample replicates across ten
`contrived (enzymatically fragmented cell-line gDNA) samples representing short variants, rearrangements, and
`copy number alterations (losses). The LoD was determined using the conservative hit rate approach for the
`majority of variants. A probit model was used when appropriate (when ≥3 dilution levels with hit rates between
`10% and 90% were observed). LoD by hit rate was defined as the mean VAF value (for short variants and
`rearrangements) or mean tumor fraction value (for copy number alterations) at the lowest dilution level tested with
`at least 95% detection across replicates. The hit rate was computed as the number of replicates with positive
`variant calls per the total number of replicates tested at each level of the targeted VAF (short variants and
`rearrangements) or tumor fraction (copy number alterations). Short variants with hit rates of at least 95% at all
`dilution levels or hit rates below 95% for all dilution levels were excluded from analysis as LoD could not be
`reliably estimated.
`
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`The median estimated LoD for CDx alterations are presented in Table 11 The median LoD for targeted short
`variant, rearrangement, and copy number alterations were consistent with the platform LoD (Table 12).
`
`
`Table 11: LoD estimation for CDx alterations
`
`Gene
`
`Alteration Subtype
`
`Number of Samples
`Evaluated
`
`Indels
`
`BRCA1
`
`Substitutions
`
`BRCA2
`
`Rearrangement1
`
`Substitutions
`
`Indels
`
`BRCA2- EDA Truncation1
`
`Copy Number Loss1
`
`Indels (exon 19 deletions)
`
`EGFR
`
`Substitutions (L858R
`substitutions)
`
`1
`
`8
`
`1
`
`17
`
`2
`
`1
`
`1
`
`2
`
`2
`
`Median LoD
`
`0.38% VAF*
`
`0.34% VAF
`
`0.87% VAF
`
`0.37% VAF
`
`0.36% VAF
`
`0.48% VAF
`
`48.1% TF
`
`0.27% VAF
`
`0.34% VAF
`
`The estimated LoDs for BRCA1 and BRCA2 subs and indels were confirmed at values higher than the LoDs established in
`Table 15 (see Precision: Reproducibility and Reproducibility section below, Tables 19 and 20 for confirmed LoD values).
`*The accuracy of %VAF/%TF have not been analytically validated.
`1The LoD for these alterations was determined using clinical specimens.
`
`
`The platform LoD for short variants, rearrangements, and copy number losses are presented in Table 12. A total
`of 864 short variants were included in the platform LoD analysis. The enhanced sensitivity region of the bait set
`contains 269 of the short variants analyzed and the standard sensitivity region of the bait set contains 595 of the
`short variants analyzed. The estimated LoD for short variants is 0.40% for the enhanced sensitivity region and
`0.82% of the standard sensitivity region. The median LoD is 30.4% tumor fraction for copy number losses.
`
`Because a major component driving the detectability of a variant is genomic context (repetitiveness of the
`reference genomic region), the LoD analysis for short variants was also evaluated within categories based on
`genomic context as summarized in Table 13.
`
`
`Table 12: LoD estimation by variant type
`
`Alteration Type
`
`Number of Variants
`in Analysis
`
`Bait Set Region
`
`Median LoD
`
`Quartile 1 to
`Quartile 3 LoD Range
`
`Short Variants
`
`Rearrangements
`
`Copy Number
`Losses
`
`269
`
`595
`
`7
`
`1
`
`2
`
`Enhanced Sensitivity
`
`0.40% VAF
`
`0.33% - 0.50% VAF
`
`Standard Sensitivity
`
`0.82% VAF
`
`0.70% - 0.98% VAF
`
`Enhanced Sensitivity
`
`0.37% VAF
`
`0.26% - 0.47% VAF
`
`Standard Sensitivity
`
`0.90% VAF
`
`NA
`
`NA
`
`30.4% TF
`
`NA
`
`VAF = variant allele frequency
`TF = tumor fraction
`*The accuracy of %VAF/%TF have not been analytically validated
`Table 13: LoD by variant subtype based on genomic context
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`Region
`
`Alteration Subtype
`
`N
`
`Minimum
`LoD
`(VAF/TF)1
`
`1st Quantile
`LoD
`(VAF/TF)1
`
`Median
`LoD
`(VAF/TF)1
`
`3rd Quantile
`LoD
`(VAF/TF)1
`
`Enhanced
`Sensitivity
`Region
`
`Short Variants: Enhanced
`Sensitivity Region Total
`
`Insertion/Deletion in non-
`repetitive region or a
`repetitive region of <=3
`base pairs
`
`Insertion/Deletion in a
`repetitive region of 4 to 6
`base pairs
`
`Insertion/Deletion in a
`repetitive region of >=7
`base pairs
`
`Substitution in a non-
`repetitive region or a
`repetitive region of <=7
`base pairs
`
`269
`
`0.20%
`
`0.33%
`
`0.40%
`
`0.50%
`
`10
`
`0.23%
`
`0.29%
`
`0.31%
`
`0.36%
`
`23
`
`0.28%
`
`0.37%
`
`0.48%
`
`0.56%
`
`6
`
`0.33%
`
`0.48%
`
`0.58%
`
`0.82%
`
`229
`
`0.20%
`
`0.33%
`
`0.39%
`
`0.49%
`
`Substitution in a repetitive
`region of >7 base pairs
`
`1
`
`0.32%
`
`0.32%
`
`0.32%
`
`0.32%
`
`Short Variants: High
`Sensitivity Region Total
`
`Insertion/Deletion in non-
`repetitive region or a
`repetitive region of <=3
`base pairs
`
`Insertion/Deletion in a
`repetitive region of 4 to 6
`base pairs
`
`595
`
`0.40%
`
`0.70%
`
`0.82%
`
`0.98%
`
`18
`
`0.46%
`
`0.68%
`
`0.87%
`
`1.00%
`
`32
`
`0.61%
`
`0.75%
`
`0.87%
`
`0.95%
`
`Insertion/Deletion in a
`repetitive region of >=7
`base pairs
`
`Substitution in a non-
`repetitive region or a
`repetitive region of <=7
`base pairs
`
`Substitution in a repetitive
`region of >7 base pairs
`
`Rearrangements
`
`11
`
`0.59%
`
`1.07%
`
`1.15%
`
`1.20%
`
`524
`
`0.40%
`
`0.70%
`
`0.81%
`
`0.96%
`
`8
`
`7
`
`0.69%
`
`0.83%
`
`0.96%
`
`1.28%
`
`0.20%
`
`0.26%
`
`0.37%
`
`0.47%
`
`Rearrangements
`
`1
`
`0.28%
`
`0.28%
`
`0.28%
`
`0.28%
`
`Rearrangements
`
`1
`
`1
`
`0.90%
`
`0.90%
`
`0.90%
`
`0.90%
`
`12.70%
`
`12.70%
`
`12.70%
`
`12.70%
`
`Standard
`Sensitivity
`Region
`
`Enhanced
`Sensitivity
`Region
`
`Enhanced/
`Standard
`Sensitivity
`Region
`
`Standard
`Sensitivity
`Region
`
`NA
`
`Copy Number Losses
`
`1VAF reported for short variant and rearrangement LoD, tumor fraction reported for copy number loss LoD.
`The accuracy of %VAF/%TF have not been analytically validated
`
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`Page 12 of 35
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`RAL-0035-01
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`Case 1:20-cv-01580-LPS Document 1-14 Filed 11/23/20 Page 14 of 36 PageID #: 503
`
`The median LoD for highly-actionable, non-CDx alterations evaluated for LoD are presented in Table 14.
`The median LoD for these targeted short variants are consistent with the platform LoD presented in Table
`12.
`
`
`
` Table 14: LoD for non-CDx alterations
`
`Gene
`
`Alteration Subtype
`
`Number of Samples
`Evaluated
`
`Median LoD*
`
`ATM
`
`BRAF
`
`KRAS
`
`MET
`
`NRAS
`
`PALB2
`
`Indels
`
`Substitutions
`
`Substitutions
`
`Indels
`
`Substitutions
`
`Indels
`
`Substitutions
`
`PIK3CA
`
`Substitutions
`
`1
`
`1
`
`2
`
`1
`
`2
`
`1
`
`1
`
`6
`
`0.51% VAF
`
`0.33% VAF
`
`0.33% VAF
`
`0.41% VAF
`
`0.42% VAF
`
`0.37% VAF
`
`0.51% VAF
`
`0.34% VAF
`
`VAF = variant allele frequency
`*The accuracy of %VAF have not been analytically validated.
`
`
`2.4 Limit of Bla