Reliable identification of gene fusions by targeted RNA sequencing · 2020-03-06 · alk camta1...

Dr Volker Endris

Institute of PathologyHeidelberg Medical FacultyHeidelberg Germany

Reliable identification of gene fusions by targeted RNA sequencing

Universitaumltsklinikum Heidelberg | Dr Volker Endris

Conflict of interests

Lecture fees from Thermo Fisher Scientific and Astra Zeneca

Adivsory Board Novartis

Speaker was provided travel and hotel support by Thermo Fisher Scientific for this presentation but no remuneration



Mitelman database cases 69174

fusions 22001 (May 23 2019)

FusionHub entries 153110


Yoshihara et al The landscape and therapeutic relevance of cancer-associated transcriptfusions

Gene fusions in solid cancer


Yoshihara et al The landscape and therapeutic relevance of cancer-associated transcript fusions

Recurrent gene fusions


Gene fusion detection

httpsglobalntrktestingcomdetecting-NTRK-gene-fusionstesting-methodologies-overview

TargetedAmpliSeqTM

TargetedHybrid Capture

TargetedAnchored Multiplex

Whole transcriptomeRNASeq


Low input 10-15ng

Easy workflow


Heidelberg Lung Cancer Diagnostics

Volckmar et al Combined targeted DNA and RNA sequencing of advanced NSCLC in routine molecular diagnostics Analysis of the first 3000 Heidelberg cases IJC 2019


NGS-based fusion detection

Lung Fusion v2

Target Partner Target Partner

ALK

EML4KIF5BKLC1HIP1TPRNPM1TFGTPM1TPM3TPM4PPFIBP1FN1SQSTM1MSNVCL RNF213NCOA1PRKAR1APTPN3CLIP4 MYH9 CLTC

ROS1CD74SDC4SLC34A2EZRTPM3 LRIG3GOPCCEP85LTFGCCDC6

RET

KIF5BCCDC6CUX1ERC1NCOA1PRKAR1A HOOK3KTN1TRIM24TRIM27TRIM33FKBP15 KIAA1468TBL1XR1AKAP13AFAP1PARG PCM1GOLGA5

MET Exon14 skipping

NTRK1

DYNC2H1CELNFASCIRF2BP2TFGSQSTM1 SSBP2CD74MPRIPTPM3TPRMIR548F1

PDGFRA SCAF11

NRG1 CD74SLC3A2 BRAF TRIM24

FGFR1 BAG4 FGFR2 CITCCAR2

FGFR3 TACC3BAIAP2L1 ERBB4 EZR

AXL MBIP EGFR EGFRRAD51PURB

Custom AmpliSeqTM Fusion Panel

Pfarr et al GCC 2016


Identified gene fusions (~ 3000 routine MDx cases)

Custom AmpliSeqTM Fusion Panel+ALK-IHC


ALK fusion variants


Christopoulos P et al (2018) EML4‐ALK fusion variant V3 is a high‐risk feature conferring accelerated metastaticspread early treatment failure and worse overall survival in ALK+ non‐small cell lung cancer Int J Cancer 142 2589-2598 doi101002ijc31275

ALK V3+ high risk variant


ALK V3+TP53+ subset

Christopoulos P et al (2018) Identification of a highly lethalV3+TP53+ subset in ALK+ lung adenocarcinoma Int J Cancer Accepted Author Manuscript doi101002ijc31893


Resistance mutations

One-stop shop approach (DNA+Fusion) gt economicaltimetissue saving


MET exon 14 skipping

c2888-20_2888-8del


Detection of METEx14 by RNAseq

0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples

AmpliSeq Fusion Panel

13 14 15


Fusion analysis beyond lung cancer

Archer Solid Tumor Panel

AKT3 EWSR1 NOTCH12 RAF1

ALK FGFR123 NRG1 RELA

ARHGAP26 FGR NTRK123 RET

AXL INSR NUMBL ROS1

BRAF MAML2 NUTM1 RSPO23

BRD34 MAST12 PDGFRAB TERT

EGFR MET PIK3CA TFE3

ERG MSMB PKN1 TFEB

ESR1 MUSK PPARG THADA

ETV1456 MYB PRKCAB TMPRSS2

Archer Sarcoma panel

ALK CAMTA1 CCNB3 CIC EPC EWSR1FKHR

FUS GLI1 HMGA2 JAZF1 MEAF6 MKL2 NCOA2

NTRK3 PDGFB PLAG1 ROS1 SS18 STAT6 TAF15

TCF12 TFE3 TFG USP6 YWHAE

Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3

ATP1B1DNAJB1 NFIBPCDHGA1QKITYK2

C11orf95ZMYND8 ETV6SPTBN1

CD74PCM1PDE7ASDC4SLC3A2TENM4VAMP2 BRD3BRD4CICWHSC1L1

ATF7IPBIN2CAPRIN1CCDC6CCDC88CCEP85LCPSF6DTD1EBF1ERC1

ETV6GIT2GOLGA4GOLGB1HIP1KANK1MPRIPMYO18ANDE1NINPEIF3EGRHL2

PTPRK ADAMTS16CCDC127GLIS3MTMR12SLC12A7TRIOTTLL7

ATAD2EZR SLC45A3TMPRSS2

AKAP12ARMT1CCDC170DAB2MTHFD1LPDE10APOLMYBYAP1 ERGETV1ETV4ETV5

ATF7IPBCRBICD2EBF1ETV6OFD1PAX5PCM1PPFIBP1SEC31ASPAG

9SSBP2STRN3TERF2TPM3TPR

Exon14_SkippingBAIAP2L1C8orf34CAPZA2DCTN1EPS15LRRFIP1OXR1P

PFIBP1PTPRZ1TFGTPRTRIM4ZKSCAN1

CHD9EIF3EPCNXHMGA2NPC2 WIPF2

ERGETV1ETV1aETV1bETV4aETV5aETV5bETV5d TBL1XR1FNDC3B

ABCB5ACACAASIC2ATAD5PSMD11 AFAP1AGBL4DAB2IPNACC2NAV1QKISLMAPSQSTM1TRIM24VCL

76 verschiedene Partner

(uaAGAP3AGKAGTRAPAKAP9AP3B1AP3B1ARMC10ATG7BAIAP2L1

VIII-

VarianteSEPT14ACADMCAND1CDK6DYMERP44GNSPDP1PSPHSEC61

GABBR2MIR143HGNUP214SDCCAG3SEC16ASNHG7AGGF1B4GALT1C9orf153CLCN6CNTLNESRP1FYCO1GOLGA4HACL1LM

NAMPRIPPAPD7PDZRN3QKISRGAP3TRAK1TRIM33BCRCDK5RAP2DIP2CETV6FIP1L1FOXP1KDRKIF5BSCAF11STRNTNK

S2AXL

AFF3 AHCYL1 BICC1 CASP7 CCAR2 CCDC6 CD44 CIT COL14A1

CREB5 CTNNB1 FAM76A KCTD1 MGEA5 NOL4 OFD1 PARK2 PDHX

PPHLN1 SHTN1 SLC45A3 SNX19 TACC3 TXLNA USP10

AES BAIAP2L1 ELAVL3 ETV6 FBXO28 JAKMIP1 TACC3

CDK4 PAX8CREB3L2TSEN2

ADAMTS16ARHGEF2BCANCD74CELCHTOPDYNC2H1EPHB2EPS15IR

F2BP2LMNAMPRIPMRPL24NFASCPPLRABGAP1LRNF213SQSTM1SS

BP2TFGTP53TPM3TPR

AKAP13BTBD1COX5AEML4ETV6FAT1HOMER1LYNRBPMS

OCP-RNA_v3Partner Partner

A2MACTG2ATICATRNL1BEND5BIRC6CADCARSCCDC88ACLIP1CLIP

4CLTCDCTN1EML4FN1GFPT1GTF2IRD1GTF3C2HIP1KANK2KCNQ5

KIF5BKLC1KTN1MCFD2MEMO1MSNMYH9NCOA1NPM1PPFIBP1P

PM1GPPP4R3BPRKAR1ARANBP2RNF213SEC31ASQSTM1STK32BST

RNTERT und andere

CCDC6 CD74 CEP85L CLIP1 ERC1 EZR GOPC HLA_A KDELR2 KIAA1598

LRIG3 MSN MYO5A NCOR2 NFKB2 PPFIBP1 PWWP2A SDC4 SLC34A2

TFG TMEM106B TPM3 YWHAE ZCCHC8

KIF5BERC1_ELKSTBL1XR1AKAPCCDC6ERC1NCOA4PCM1HOOK3KIA

A1468(RFG9)FKBP15GOLGA5PRKAR1ATRIM24TRIM33TRIM27CUX1

SPECC1LKTN1

BAG4 ERLIN2 ADAM32 PLAG1 TACC1 ZNF703 BCR CNTRL CUX1

FGFR1OP LRRFIP1 MYO18A TRIM24 ZMYM2 TPR FGFR1OP2 WHSC1L1

FN1 SQSTM1 CPSF6 ERVK3_1 RANBP2



Assay characteristics

Archerreg FusionPlexreg Solid Tumor (AMP)

Oncominetrade Comprehensive Assay v3 (OCA)

Sample input 200 ng RNA8 x 7microm

20 ng RNA4 x 7microm

Detection method Anchored multiplex PCR Targeted NGS amplicon based

Genetic regions 53 gt1000 specific fusions

Platform Ion Torrenttrade or Illuminareg Ion Torrenttrade

Fusion detection All fusions of the included genes in parallel

Many genes in parallel


ESR1QKI (E2Q5)

RNF130SEPT14 (R3S10)

TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)

Identification of previously unknown gene fusionsis possible if primers for the fused genes are presentin the panel

Non-targeted detection of gene fusions OCAv3


Identified gene fusions


NTRK gene fusions are rare but recurrent oncogenic drivers

AAAA

Promoter

5rsquo partner TRK kinase domain

5rsquo partner kinase domain

NTRK123

LBD

ERK

AKT

Amino terminal dimerization domain

TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP

from Ulrik Lassen presented ESMO2018

NTRK gene fusions


NTRK fusions across entities


1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2

Diversity of cancers treated

ThyroidInfantile fibrosarcoma

Salivary gland

LungColon

Melanoma

Gastrointestinal stromal tumor

Cholangiocarcinoma

Bone sarcoma

Breast

Appendix PancreasCongenital mesoblastic nephroma

Unknown primary Spindle cell sarcomaInflammatory myofibroblastic tumor

Sarcoma NOS

Peripheral nerve sheath

Myopericytoma

Infantile myofibromatosis

Lipofibromatosis

Small round cell sarcoma

Stromal sarcoma

Inflammatory myofibroblastic kidney tumor

Not determined

Primary dataset (n=55) Supplementary dataset (n=67)

Subtypes of soft tissue sarcoma

NOS not otherwise specified


NTRK fusions are therapeutically responsive

-100

-90-80-70-60-50-40-30-20-10

01020304050 Infantile fibrosarcoma

Soft tissue sarcomaThyroidSalivary gland

MelanomaBreast

LungAppendix

GIST

ColonPancreasCholangiocarcinoma

Integrated dataset Larotrectinib is efficacious regardless of tumor type

Congenital mesoblastic nephromaUnknown primaryBone sarcoma

Max

imu

m c

han

ge in

tu

mo

r si

ze (

)

DaggerIncludes 9 unconfirmed PRs pending confirmation does not include 13 patients continuing on study and awaiting initial response assessment Patient had TRKC solvent front resistance mutation (G623R) at baseline due to prior therapy Surgical CR daggerRECIST 11 Note Two patients not shown here These patients discontinued treatment prior to any post-baseline tumor measurementsCR complete response ORR objective response rate PR partial response

Investigator response assessments as of 30 July 2018

IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17


Testing algorithm for TRK fusion cancer

Penault-Llorca et al J Clin Pathol 2019 Jul 72(7) 460ndash467


Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion

AmpliSeqTM Fusion detection is a robust high-throughputtechnology that allows sensitive and specific detection of genefusions with as little as 10ng RNA


Thank youMolDia-Team HeidelbergRoland PenzelJan BudcziesAnna-Lena VolckmarMartina KirchnerRegine BrandtOlaf NeumannSuranand TallaEugen RempelDaniel KazdalAlbrecht StenzingerPeter Schirmacher

Thoracic HospitalPetros ChristopoulosMichael Thomas

NCTTilmann BochtlerAlwin Kraumlmer


Thermo Fisher Scientific and its affiliates are not endorsing recommending

or promoting any use or application of Thermo Fisher Scientific products

presented by third parties during this seminar Information and materials

presented or provided by third parties are provided as-is and without

warranty of any kind including regarding intellectual property rights and

reported results Parties presenting images text and material represent they have the rights to do so


FISH-negative RET cases

RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24

KIF5B-RET (Fall 3) CCDC6-RET (Fall 9)

KIF5B-RET (K15R12)


Conflict of interests

Lecture fees from Thermo Fisher Scientific and Astra Zeneca

Adivsory Board Novartis

Speaker was provided travel and hotel support by Thermo Fisher Scientific for this presentation but no remuneration




fusions 22001 (May 23 2019)











TargetedAmpliSeqTM





Low input 10-15ng

Easy workflow






Lung Fusion v2


ALK



RET


MET Exon14 skipping

NTRK1


PDGFRA SCAF11











ALK fusion variants





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)




fusions 22001 (May 23 2019)











TargetedAmpliSeqTM





Low input 10-15ng

Easy workflow






Lung Fusion v2


ALK



RET


MET Exon14 skipping

NTRK1


PDGFRA SCAF11











ALK fusion variants





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)










TargetedAmpliSeqTM





Low input 10-15ng

Easy workflow






Lung Fusion v2


ALK



RET


MET Exon14 skipping

NTRK1


PDGFRA SCAF11











ALK fusion variants





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)


Low input 10-15ng

Easy workflow






Lung Fusion v2


ALK



RET


MET Exon14 skipping

NTRK1


PDGFRA SCAF11











ALK fusion variants





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)






Lung Fusion v2


ALK



RET


MET Exon14 skipping

NTRK1


PDGFRA SCAF11











ALK fusion variants





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)





ALK fusion variants





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)





ALK V3+TP53+ subset







c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)






c2888-20_2888-8del



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)



0

20000

40000

60000

80000

100000

120000

140000

160000

1 10 100 1000

Re

ad c

ou

nts

Tested samples


13 14 15







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)







AXL INSR NUMBL ROS1




ERG MSMB PKN1 TFEB








Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1

















ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)














ESR1QKI (E2Q5)


TRIM24BRAF (T9B11)

BRD4NUTM1 (B14N2)







AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)





AAAA

Promoter



NTRK123

LBD

ERK

AKT


TRK kinase domain

Tyr

Tyr

TRK kinase domain

Tyr

Tyr

PP

PP


NTRK gene fusions




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)




1916

10

107

6 4 3 33

3 31

1

1

1

1

1

1

22

13

9

7

77 5 5

44

4

4

2

2

2

2

2



Salivary gland

LungColon

Melanoma


Cholangiocarcinoma

Bone sarcoma

Breast



Sarcoma NOS


Myopericytoma


Lipofibromatosis


Stromal sarcoma


Not determined






-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)



-100

-90-80-70-60-50-40-30-20-10



MelanomaBreast

LungAppendix

GIST




Max

imu

m c

han

ge in

tu

mo

r si

ze (

)



IntegratedDagger

(n=109)

ORR (95 CI)dagger 81 (72‒88)

Best responsedagger

PR 63

CR 17





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)





Target Target

ALK ROS1

RET FGFR1

FGFR2 FGFR3

UBA1 PPARG

NTRK1 NTRK3

BRAF EGFR

NOTCH1 RAF1

PDGFRA MBIP

RAD51B ERBB2

TMPRSS2 PIK3CA

NF1 NTRK2

ERBB4 ERG

ESR1 TMPRSS2

JAK2 MET

NRG1 NUTM1

PDGFRB RSPO2

RSPO3 TERT

PRKACAB MYB

RELA FLT3


















VIII-




S2AXL








BP2TFGTP53TPM3TPR







RNTERT und andere






SPECC1LKTN1





Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)


Conclusion















RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)














RET

KIF5B

Chr10

RETEx 1 ndash 1524 Ex 1112 - 24


KIF5B-RET (K15R12)

Reliable identification of gene fusions by targeted RNA sequencing · 2020-03-06 · alk camta1...

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Transcript of Reliable identification of gene fusions by targeted RNA sequencing · 2020-03-06 · alk camta1...