savannah, april 2019
TRANSCRIPT
10/16/2019
1
Ancillary Tests for Melanoma Diagnosis
KJ Busam, MD, MSKCCNew York, NY
Savannah, April 2019
NO CONFLICTS OF INTEREST
The Rise of Molecular Tests
DiagnosisCytogenetics
Gene ExpressionGene Fusions
Mutations
Prognosis Gene Expression
TargetedTherapy
Mutation Analysis
TEST TESTING SITE COST
Gene expression, adhesive patch
Commercial • $1100 ($50 co-pay if paid within 30 days)
• $249 self pay
FISH Commercial • $2400/4 probes• $600/single probe • Technical only: $1760/4 probes• Technical only: $440/single probe
Academic • $1200/4 probes• $600/single probe
Array CGH Academic • $1800
Gene expression, diagnosis
Commercial • Internal research not yet billed to patients or insurance
Gene expression, prognosis
Commercial • $7000
Sequencing, next generation
Commercial • $5800
The Rise of Molecular Tests
1
2
3
10/16/2019
2
Lesson # 1
Molecular test can be essential
Case 1
41 F with h/o melanoma andnodule in lung
S100P Pos. also:- Melan-A- HMB-45
Reported Diagnosis
4
5
6
10/16/2019
3
- Metastatic tumor submitted for NGS to check for targetable mutation- NGS – TEST RESULT:
Sequence Analysis
Pathologic – Molecular Correlation
• Tumor with melanocytic differentiation
• Fusion of EWSR1 with a member of the CAMPResponse Element Binding protein family
– ATF1
– CREB
– CREM (CRE-Modulator)
CLEAR CELL SARCOMA
Pathology Report of Primary
7
8
9
10/16/2019
4
S100P Pos alsofor:- HMB-45- Melan-A
Primary Skin Tumor
10
11
12
10/16/2019
5
Identical EWSR1-CREM Fusion in Primary and Met
Molecular Analysis of Primary Tumor
Revised and Final Diagnosis
Clear Cell Sarcoma Metastatic to Lung
13
14
15
10/16/2019
6
Is CCS a Melanoma?
16
17
18
10/16/2019
7
CCS clusters with melanoma
J Clin Oncol 2003; 21:1775-81
CCS = variant of melanoma
19
20
21
10/16/2019
8
87 yo M with groin mass
Case 2
IHC-Results
• Pos for CD56
• Loss of H3K27me3
• Neg for S100, Sox10
Case 2
22
23
24
10/16/2019
9
87 yo M with groin mass
Reported as “MPNST”
Case 2
NRASQ61R
NRASQ61R
Melanoma of right calf
2 years earlier…Case 2
25
26
27
10/16/2019
10
Mutation Analysis of Groin Mass
Case 2
Revised Diagnosis: Metastatic Melanoma
• Lymph node involvement by tumor
• Prior melanoma at site draining to positive node
• Same mutation in melanoma as in undifferentiated tumor
• Undifferentiated tumor’s mutation profile shows UV signature
Case 2
28
29
30
10/16/2019
11
Critical Value of Mutation Analysis
• Diagnosis of Clear Cell Sarcoma
• Diagnosis of Undifferentiated Melanoma
• Diagnosis of Melanotic Schwannian Tumor
• Determining the Relationship of Multiple Melanocytic Tumors
Nevi n=54Melanoma n=133
Cytogenetic Analysis for the Distinction of Melanoma from Melanocytic Nevi
Bastian et al Am J Pathol 2003
Benign Malignant
31
32
33
10/16/2019
12
Spitzoid Melanoma of Childhood
Loss on chromosome 9
p16
Homozygous9p21 deletion
Metastatic MelanomaAm J Surg Pathol. 2013;37:676-84.
2F, shoulder
Case 3
3M, Cheek
Case 4
What is Your Diagnosis?
What is Your Diagnosis?
3M, cheek
34
35
36
10/16/2019
13
Cytogenetics for Diagnosis
• Can be valuable for difficult cases
– Nevoid melanoma
– Malignant Spitz tumor or spitzoid melanoma
– Blue melanoma
• Often used for “hand-holding”
Lesson # 2
Molecular test may not be essential, but results can come in handy
37
38
39
10/16/2019
14
• Sclerosing BN vs Desmoplastic Melanoma
• BN vs Pigmented Epithelioid Melanocytoma
• True Spitz vs Spitzoid
Mutation Analysis for Diagnosis
Sensitivity- 87%Specificity- 96%
Is FISH really necessary?
14 M, foot lesionCBN with rate mitosis
40
41
42
10/16/2019
15
Is FISH really necessary?
70 F with new pigmented lesion at site of melanoma scar
Cytogenetics– a “Game Changer”?
42 yo womanLt foot
43
44
45
10/16/2019
16
Unnecessary Testing
Lesson # 3
Molecular findings help clarify tumor classifications
Molecular Classification of Spitz Lesions
46
47
48
10/16/2019
17
BAP1-Deficient Epithelioid Melanocytic Nevus/Tumor
Wiesner et al. Am J Surg Path 2012;36:818-30.
49
50
51
10/16/2019
18
Deep Penetrating Nevus (DPN)
B L U E Melanomas
• BLue nevus-related
• Uveal melanoma Equivalent– No association with epithelia
– Morphologic spectrum
– Driver mutation
– Prognostic factors
– Behavior
What is Your Diagnosis?
52
53
54
10/16/2019
19
CBN vs DPN vs PEM?
Diagnosis: Blue Nevus
Mutation: CYSLTR2
What level of detail is needed for clinical care ?
How much does a “benign melanocytic tumor” need to be subclassified?
55
56
57
10/16/2019
20
Lesson # 4
Molecular tests results can be misleading
18 F, ear canalOrdinary Melanocytic Nevus with Positive Melanoma FISH Test
18 F, ear canalFISH:-Gain of 6p-Loss of 6q
“Positive” FISH Test
58
59
60
10/16/2019
21
False-positive Cytogenetic Result
• FISH:
– Enumeration error
• “Cherry picking”
• Truncated nuclei
– Tetraploidy
• CGH and FISH:
– A benign or indolent “borderline” tumor may have genomic aberrations
False-negative Cytogenetic Results
• Technical
– FISH: Aberrations on chromosomes not targeted by FISH probes
– CGH: Tumor diluted by normal tissue
• Biological
– Melanoma without detectable genomic aberrations
61
62
63
10/16/2019
22
TERT promoter Mutations for the Diagnosis of Melanoma
• 86 primary cutaneous melanomas
• 72melanocytic nevi
• Sensitivity for melanoma: 78%
• Specificity for melanoma: 98%
Ordinary Melanocytic Nevus with 124C>T TERT Mutation
Molecular findings can be a useful piece of the diagnostic puzzle
64
65
66
10/16/2019
23
What is Your Diagnosis?
50 M, Thigh
Submitted by clinician as “r/o irritated SK”
Nevus or Melanoma?
Pathology Report
67
68
69
10/16/2019
24
NTRK1
What to do, if you consider a Spitz tumor?
Molecular Tests
• Archer Fusion: LMNA- NTRK1
• SNP Array: No unbalanced genomic aberration
Diagnosis: Spitz Nevus
50 M, Thigh
Submitted by clinician as “r/o irritated SK”
70
71
72
10/16/2019
25
PRAME
Diagnosis: Nevoid Melanoma
Several unbalancedgenomic aberrations
PRAME
• PReferentially expressed Antigen in Melanoma
• Cancer Testis Antigen
PRAME
Gene Expression Data
• PRAME expressed in 90% of melanomas
• PRAME also expressed in other cancers– Sarcomas, carcinomas, neuroblastoma, germ cell
tumors, leukemia, basal cell carcinoma
• PRAME expressed in normal tissue– Testis
– Ovary, placenta, Endometrium
73
74
75
10/16/2019
26
PRAME Expression for DiagnosisAdhesive Patch Test
PRAME Expression for Diagnosis
Am J Surg Pathol 2018;42: 1456 - 1465
76
77
78
10/16/2019
27
PRAME Expression by IHC
• Metastatic Melanoma (n=100) 87% POS
• Primary Melanoma (n= 155) 83% POS
• Melanocytic Nevi (n= 145) 14% POS
PRAME in Primary Melanoma
PRAME
PRAME Expression by IHC
• Metastatic Melanoma (n=100)
• Primary Melanoma (n= 155) 83% POS
• Melanocytic Nevi (n= 145)
79
80
81
10/16/2019
28
PRAME in Primary Melanoma
PRAME
Clinical Utility of PRAME IHC
• Diagnosis of melanoma– Nevus vs nevoid/spitzoid melanoma– Nodal nevus versus metastatic melanoma– Microstaging of melanoma associated with a nevus
• Margin assessment of melanoma– Acral and LM melanoma in situ
Molecular Studies for Treatment
• NGS for Mutation Burden– Immunotherapy
• Mutations– BRAFV600E, RASQ61R
• Gene-Fusions– ALK, NTRK, ROS1, MET
Oncogenic
Targets
82
83
84
10/16/2019
29
VE1 for Detection of BRAFV600E
Limitation of IHC (VE1)
• Recognizes only BRAFV600E
• A negative test result does not exclude other targetable BRAF mutations
• IHC for RAS helpful, since if positive other BRAF mutations are unlikely
Rapid BRAF Test
• Recognizes only BRAFV600E
• A negative test result does not exclude other targetable BRAF mutations
• IHC for RAS helpful, since if positive other BRAF mutations are unlikely
85
86
87
10/16/2019
30
KINASE FUSIONS
IHC of Kinases – Relevance
• Surrogate for presence of kinase fusion
• Classification of Spitz vs Spitz-like
• Potential Treatment Option
ALK-Positive Melanomas
88
89
90
10/16/2019
31
ALKATI Isoform in Metastatic Melanoma
Nature 2015 (in press)
Anal Melanoma with NTRK Fusion
NTRK
Am J Surg Pathol2018
TRAF2-NTRK2Fusion
47F with peri-anal melanomametastatic to LN
Targeting TRK Fusions
91
92
93
10/16/2019
32
Metastatic Cutaneous Melanomawith STX7-ROS1 Fusion
• Metastases to mediastinum, intestine
• BRAF wild-type
• POD on nivolumab + ipilimumab
58 F with stage IV melanoma; primary melanoma in 1993
Metastatic Melanoma ROS1 Fusion
Sox10
ROS1
ROS1
Radiographic CR ongoing at 20+ weeks
PATHOLOGY OF CR: NECROTIC TUMOR NODULE – NO VIABLE TUMOR, C/W TREATMENT EFFECT
Metastatic Melanoma with ROS1 Fusion
94
95
96
10/16/2019
33
Pathology for Treatment Selection
• Mutations:
– BRAFV600
– Mutation Burden
• Gene Fusions (Alk, Ntrk, Ros1)
Acknowledgements
• Members of the IHC Lab
– A Jungbluth
– D Frosina
• Members of the Dermpath team
– C Lezcano
– T Hollmann
– M Pulitzer
97
98