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Emerging Molecular Diagnostic
Tests and Therapies for Melanoma::
From Research Benches to
Clinical TrenchesClinical Trenches
Aleodor (Doru) Andea, MD
Associate Professor of Pathology and Dermatology
Director of Dermatopathology Section
University of Alabama at Birmingham
Birmingham, USA
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Disclosure information
I have no financial or industrial affiliation to
disclose.
Aleodor A Andea
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Overview
1. Problems in the diagnosis
2. Molecular alterations (with implications
for diagnosis or treatment)
3. Diagnostic assays3. Diagnostic assays
4. New therapies
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Melanoma diagnosis
1.Size
2.Symmetry
3.Circumscription
5.Nested architecture
NEVI MELANOMA
Small Large
Symmetric Asymmetric
Well-circumscribed Poorly-circumscribed
Predominantly nested Predominantly single cells
4.EpidermisNot affected Consumption, Ulceration
7.Pagetoid spread
8.Confluent growth at DEJ
10.Maturation
6.Nests uniformity
11.Cytologic atypia
12.Mitoses in dermis
Uniform Irregular
No Prominent
No Present
Present Absent
Absent Severe
Absent Present, Atypical
9.Dermal patternNested, Infiltrative Sheetlike, Expansile
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• Age
• Site
• History of trauma
Melanoma diagnosis
Factors that influence interpretation of histologic
parameters
• History of trauma
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2 y/o girl with lesion on the rt knee
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Dermatopathology diagnosis
Spitz tumor of uncertain malignant
potential
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Dermatopathology diagnosis
Spitz tumor of uncertain malignant
potential
Diagnosis 1 Diagnosis 2Diagnosis 1
Mitotically active
Spitz nevus
Diagnosis 2
Spitzoid melanoma
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• ~1-2 million bx/ year in US to rule out
melanoma
• ~3%-6% of bx are melanomas
• In most cases dx can be made on • In most cases dx can be made on
histopathology but a small proportion
have ambiguous histology (Q.)
• 2.3%-25% of cases with diagnostic
discrepancies (McGinnin et al, Arch Dermatol 2002, Corona et al, J Clin Oncol 1996, Farmer et al, Hum Pathol 1996, Lodha et al J Cutan Pathol 2008)
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• Ambiguous lesions with overlapping
criteria
– Severely atypical/ dysplastic nevus vs LM
or SSM
– Nevi of special site vs LM or SSM
– Spitz nevus vs spitzoid melanoma– Spitz nevus vs spitzoid melanoma
– Atypical blue nevi vs melanoma
– Proliferative nodules vs melanoma
– Nevoid melanoma
• Inter-observer variability
Zembowicz and Scolyer. Arch of Pathol and Lab Med. 2011; 135
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57 borderline lesion with clinical follow-up
• 17-benign• 17-benign
• 26-melanoma
• 14-borderline
Panel performance:
• 73% sensitivity
• 47% specificity
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Need for a better test
• Prevent under/overtreatment of patients
• Reduce medical costs associated with unnecessary treatmentunnecessary treatment
• Impact positively on patient care
• Most common reason for medical malpractice in pathology
Kornstein, et al. Arch Pathol and Lab Med, 2007
Gerami, et al, Am J Surg Pathol, 2009
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Danube River, Romania
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Molecular alterations in
melanocytic neoplasms
• MAP kinase alterations
• Replicative senescence in melanocytic
lesionslesions
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GROWTH
FACTORS
RAS
RAF
CYTOPLASM
H-Ras, K-Ras, N-Ras
A-Raf, B-Raf, C-Raf1
MAP KinaseKIT, EGFR
MEK
ERK
PROLIFERATION
CYTOPLASM
NUCLEUS
MEK-1, MEK-2
ERK-1, ERK-2
CCND1
CDK4/6
p16
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GROWTH
FACTORS
RAS
RAF
CYTOPLASM
KIT mutations in 14-19% of melanomas
MAP Kinase pathway activation in
melanocytic lesions
MEK
ERK
CYTOPLASM
NUCLEUS
Curtin JA et al. J. Clin. Oncol. 2006; 24; 4340–6
Beadling C et al. Clin Cancer Res 2008; 14:6821-8
PROLIFERATIONCCND1
CDK4/6
p16
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GROWTH
FACTORS
NRAS
RAF
CYTOPLASM
10-22% of melanomas
80% large congenital nevi
MAP Kinase pathway activation in
melanocytic lesions
MEK
ERK
CYTOPLASM
NUCLEUS
Curtin JA, et al. N Engl J Med. 2005 Nov 17;353(20):2135-47
PROLIFERATIONCCND1
CDK4/6
p16
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GROWTH
FACTORS
HRAS
RAF
CYTOPLASM
27-29% of Spitz’s nevi
MAP Kinase pathway activation in
melanocytic lesions
MEK
ERK
CYTOPLASM
NUCLEUS
Bastian B et al, Am J Pathol 2000
Van Dijk M et al. Am J Surg Pathol 2005; 29:1145-51
PROLIFERATIONCCND1
CDK4/6
p16
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GROWTH
FACTORS
RAS
B-RAF
CYTOPLASM
MAP Kinase pathway activation in
melanocytic lesions
40-60% of melanomas (Q.)
80% of nevi
MEK
ERK
CYTOPLASM
NUCLEUS
Davies et al, Nature 2002, Marcia et al. Cancer Res 2002, Curtin et al, N Engl J Med 2005, Pollock et al., Nat Genet 2002
PROLIFERATIONCCND1
CDK4/6
p16
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GROWTH
FACTORS
RAS
B-RAF
CYTOPLASM
MAP Kinase pathway activation in
melanocytic lesions
GNAQ
PKC
Guanine nucleotide-binding protein
G(q) subunit alpha
•Blue nevi: 80%
•Uveal melanoma: 50%
MEK
ERK
CYTOPLASM
NUCLEUS
Van Raamsdonk C et al. Nature 2009 Ian 29; 457:599-603
PROLIFERATIONCCND1
CDK4/6
p16
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Nevus type KIT NRAS HRAS KRAS BRAF GNAQ
Congenital NA 70-81% 0 NA 0-
88%
0
Acquired NA NA 0 NA 70-
88%
0
Clark’s NA NA 0 NA 52-
80%
NA
Spitz NA 0% 27- NA 0% 0
Genetic alterations in nevi
Spitz NA 0% 27-
29%
NA 0% 0
Blue nevus NA 0-5% NA 0-15% 0-12% 40-
83%
Curtin JA, et al. N Engl J Med. 2005 Nov 17;353(20):2135-47, Maldonado JL et al. J Natl
Cancer Inst. 2003 Dec 17;95(24):1878-90.
Satzger I et al. Br J Cancer. 2008 Dec 16; 99(12):2065-9
Emley A et al. Hum Pathol. 2011 Jan; 42:136-40
Van Raamsdonk C et al. Nature 2009 Ian 29; 457:599-603
Yazdi et al. J Invest Dermatol. 2003 Nov; 121:1160-2
Bastian B et al, Am J Pathol 2000
Saldanha et al. Int J Cancer 2004 Sep 20;111:705-10
Van Dijk M et al. Am J Surg Pathol 2005; 29:1145-51
Bauer et al. J Invest Dermatol 2007; 127:179-182
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Melanoma type KIT NRAS HRAS KRAS BRAF GNAQ
MM on skin
without chronic
sun damage
0% 22% 0 0% 59% 0
MM on skin with
chronic sun
damage
16% 15% 0 0% 11% 15%
Genetic alterations in melanomas
damage
Acral 10-23% 10% 0 0% 23% 0%
Mucosal
melanoma
15-21% 5% 0 0% 11% 0
Uveal melanoma 0% 0% NA 0% 0% 50%
Malignant BN NA NA NA NA NA 46%
Curtin JA, et al. N Engl J Med. 2005 Nov 17;353(20):2135-47,
Curtin JA et al. J. Clin. Oncol. 2006; 24; 4340–6
Maldonado JL J Natl Cancer Inst. 2003 Dec 17;95(24):1878-90.
Satzger I, Br J Cancer. 2008 Dec 16;99(12):2065-9
Zuidervaart, W et al. Br J Cancer 2005 Jun 6; 92:2032-8
Beadling C et al. Clin Cancer Res 2008; 14:6821-8
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HRAS NRAS BRAF
Spitzoid melanoma 0 19% 64%
AtypicalSpitz nevus 14% 0% 0%
Spitz nevus 29% 0% 0%
Practical implication for DX
van Dijk MC et al. Am J Surg Pathol. 2005 Sep;29:1145-51.
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Orthodox Cathedral, Timisoara, Romania
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Replicative senescence in
melanocytic lesions
• Cells exit cell cycle after a number of
divisions
• Chromosomes are protected by • Chromosomes are protected by
telomeres and the enzyme telomerase
Greider and Blackburn EH, Nature 1989
Szostak et al, Cell 1989
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Chromosome
Chromosome
Chromosome
P53, RB
T T T T
T T T
T T
p53, RB,
p16
Mitosis
MitosisCheckpoint
Cell cycle arrest
(Senescence)
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Chromosome
Chromosome
Chromosome
Chromosome
T T T T
T T T
T T
T
p53, RB,
p16
P53, RB
Mitosis
Mitosis
Mitosis
Chromosome
Chromosome
T
Mitosis
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Chromosome
Chromosome
Chromosome
Chromosome
T T T T
T T T
T T
T
p53, RB,
p16
P53, RB
Mitosis
Mitosis
Mitosis
Chromosome
Chromosome Cell Crisis
DNA breaks, fusions
T
MitosisMitosis
•Gross chromosomal
abnormalities
•Increase apoptosis
Cell Death
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Chromosome
Chromosome
Chromosome
T T T T
T T T
T T
p53, RB,
p16
Mitosis
Mitosis
Nevi MAPK
-B-RAF
-N-RAS
-H-RAS
-GNAQ
Cell cycle arrest
(“Oncogene-
Induced”
Senescence)
No chromosomal aberrations
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Chromosome
Chromosome
Chromosome
Chromosome
T T T T
T T T
T T
T
p53, RB,
p16
P53, RB
Mitosis
Mitosis
Mitosis
Melanoma
Cell cycle arrest
(Senescence)
MAPK
-B-RAF
-N-RAS
-H-RAS
-GNAQ
p16 inactivation:
~50% of MM
Chromosome
Chromosome Cell Crisis
DNA breaks, fusions
T
MitosisMitosis
Cell Death
-Re-stabilize
telomeres
-growth advantage
Melanoma
Gross chromosomal abnormalities
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Parliament House, Bucharest, Romania
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Molecular tests differentiating
melanoma from nevi
• IHC
• CGH
• FISH
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Ki-67Nevus
(<5%)
Melanoma (Q.)
(>5%)
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HMB-45/ Cyclin D1
Nevus (Q.) Melanoma
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HMB-45 problemsCombined Nevus Melanoma
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p16
Nevus Melanoma
MAPK
activation
P16
inactivation
(and others)
PrecursorUnchecked
division/ growthCell crisis
Reactive p16
overexpresionNo p16
expresion
~50%
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p16
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p16Spitz Nevus Melanoma
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p16Spitz Nevus Melanoma
Stefanaki et al, J Am Acad Dermatol 2007
Positive in
100% of Spitz
nevi
Negative in 32-50%
of melanomas
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Comparative Genomic
Hybridization
• Screens the entire genome for gains
and losses in DNA material in one
experiment (Q.)experiment (Q.)
• Variants:
– Conventional CGH
– Array based CGH
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Conventional CGH
www.empiregenomics.com
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Bauer, J. and Bastian, B. Dermatologic Therapy, Vol 19, 2006, 40-49.
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The ratio of green:red signals along each chromosome is determined
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Array CGH
• Arrays of genomic bacterial artificial
chromosome (BAC) clones or
oligonucleotides
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CGH in Melanocytic lesions
• 54 benign nevi – 27 Spitz nevi
– 19 Blue nevi
– 7 Congenital nevi
• 132 MM – 22 Acral location
– 108 non-Acral
Bastian B et al, Am J Pathol 2003
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Gains: 6p Losses: Gains: 11p: 11% Losses: Gains: 6p
1q
7p
7q
8q
17q
20q
Losses:
9p
9q
10q
10p
6q
11q
Gains: 11p: 11%
7q: 2%
Losses:
0%
•The 7 cases were all Spitz
nevi (no progression to MM
at 7 yrs FU)
Bastian B et al, Am J Pathol 2003
96% of cases
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Conclusion
• CGH will show multiple gains and
losses of DNA material in the majority
of cases (Q.).
• Potential diagnostic test for ambiguous • Potential diagnostic test for ambiguous
melanocytic lesions.
• Sensitivity 96%
• Specificity 98%
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Chromosomal Aberrations in
Melanocytic Lesions
• Spitz nevi:
– no abnormalities
– gains on 11p (12-18%)
• Congenital nevi: no abnormalities
• Proliferative nodules: no abnormalities or whole chromosomal gains or losses
• Cellular blue nevi: no abnormalitiesBastian B et al, Am J Pathol 2000
Bastian B et al, J Invest Pathol 1999
Bastian B et al, Am J Pathol 2002Maize Jr, JC et al, Am J Surg Pathol 2005
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Disadvantages of CGH
• Requires 30-50% pure tumor cells
• Does not allow histologic correlation
• Cannot detect tumor subpopulations• Cannot detect tumor subpopulations
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FISH
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• 1q31 (COX2)
• 4q12 (KIT)
• 7q34 (BRAF)
• 6p35 (RREB1)
• 6q23 (MYB1)
• 6 cen
• 7 cen
• 9p31 (p16)
CGH date from
melanomas
• 9p31 (p16)
• 10 cen
• 11q13 (CCND1)
• 17q25 (TK1)
• 17q21 (RARA)
• 17 cen
• 20q13 (ZNF217)
Gerami et al. Am J Surg Pathol 2009
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• Training cohort 301 melanocytic tumors
– 148 melanomas
– 153 nevi
• Validation cohort
– Unequivocal lesions:
• 83 melanomas
• 86 nevi
– 27 ambiguous cases with clinical follow-up
• 6 cases developed metastases
• 21 free of disease at > 5 years follow-up
Gerami et al. Am J Surg Pathol 2009
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Gerami et al. Am J Surg Pathol 2009
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Gerami et al. Am J Surg Pathol 2009
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Gerami et al. Am J Surg Pathol 2009
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11q13
CCND1
6p25
RREB1
6cen 6q23
MYB1
Gain Gain Loss
>38% nuclei
contain >2
signals for
11q13
>29% nuclei
contain >2
signals for 6p25
>55% nuclei
with >1 ratio of
6p25 / 6cen
>40% nuclei
with <1 ratio of
6q23 / 6cen
CCND1 RREB1 MYB1
OR OR
OR
Gerami et al. Am J Surg Pathol 2009
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• Validation cohort:
– 72 of 83 melanomas FISH positive
• 87% sensitivity
– 82 of 86 nevi FISH negative
• 95% specificity
• 4 nevi were FISH positive• 4 nevi were FISH positive
Gerami et al. Am J Surg Pathol 2009
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• 27 cases with indeterminate histology
– 6 developed metastases, all FISH positive
• 100% sensitivity
– 21 disease free at > 5 years, 15 FISH
negative
• 71% specificity
Gerami et al. Am J Surg Pathol 2009
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CCND1 –green
RREB –red
MYB –gold
CEP6 -aqua
20 Nevi 20 Melanomas
19/20 negative
Specificity 95%
18/20 positive
Sensitivity 90%
Morey et al, Pathology 2009
MYB lossNo alterations
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Superficial
Spreading
N=70
Lentigo
Maligna
N=28
Nodular
N=22
Acral
lentiginous
N=3
DM
N=15
Sensitivity of FISH in melanoma subtypes
Sensitivity 81% 82% 91% 100% 47%
Most
common
alteration
6p25 gain
(RREB1)
73%
6p25 gain
(RREB1)
68%
6p25 gain
(RREB1)
82%
6p25 gain
(RREB1)
100%
6p25 gain
(RREB1)
86%
Gerami et al. Arch Dermatol 2010; 146:273-8
Gerami et al. J Cutan Pathol 2011; 38: 329–334
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22 total lesions
- Sensitivity: 60%
FISH positive
- Sensitivity: 60%
- Specificity: 50%
12 ambiguous lesions
FISH worked in 8 cases
-5 malignant (3 FISH
positive)
-3 benign (1 FISH negative)
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Summary of
sensitivity/specificity estimates
1 2 3 4 5 6 7
Study sample 169 233 40 43 10 324 324
Sensitivity 87% 83% 90% 85% 42% 84%
(75)
64%
Specificity 95% 94% 95% 90% 100% 95% 98%
1. Gerami et al. Am J Surg Pathol 2009
2. Gerami et al. Arch Dermatol 2010; 146:273-8
3. Morey et al, Pathology 2009
4. Vergier et al. Mod Pathol 2011;24:613-23
5. Gaiser et al, Mod Pathol 2010; 23:413-9
6. NeoGenomics validation set
7. NeoGenomics validation set with original cutoff criteria
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Summary of sensitivity
estimates on ambiguous lesions
1 2 3
Study sample 27 113 8
Sensitivity 100% 43% 60%
Specificity 71% 80% 33%
1. Gerami et al. Am J Surg Pathol 2009
2. Vergier et al. Mod Pathol 2011;24:613-23
3. Gaiser et al, Mod Pathol 2010; 23:413-9
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Reasons for False
Positives/Negative• Technical problems
• Lack of experience
• Tetraploidy in Spitz nevi (5-10%)
• Detection of non-significant clones • Detection of non-significant clones
• Signal cutoffsProbe Criterion Original Cutoff Neo
Genomics
Cutoff
RREB1 >2 RREB1 >29 >16
RREB1 RREB1>CEP6 >55 >53
MYB MYB<CEP6 >40 >42
CCND1 >2 CCND1 >38 >19
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Other potential applications
• “Nevoid” melanomas vs “Mitotically
active” nevi
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Mitotically active
nevus
N=10
Nevoid melanoma
N=10
Gerami et al. Am J Surg Pathol 2009
10/10 cases FISH negative 10/10 cases FISH positive
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Other potential applications
• “Nevoid” melanomas vs “Mitotically
active” nevi
• Intranodal nevus vs Metastatic
MelanomaMelanoma
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Metastatic Nodal nevusMetastatic
melanoma
N=24
Nodal nevus
N=17
FISH status 20 cases FISH
positive
16 cases FISH
negative
Sensitivity
83%
Specificity
94%
Scott et al. Am J Surg Pathol 2010
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Other potential applications
• “Nevoid” melanomas vs “Mitotically
active” nevi
• Intranodal nevus vs Metastatic
MelanomaMelanoma
• Blue nevus vs Blue nevus-like
metastatic melanoma
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Blue nevus-like
metastatic
melanoma
N=10
Blue nevus
N=10
FISH status 9 cases FISH
positive
10 cases FISH
negative
Sensitivity
90%
Specificity
100%
Pouryazdanparast et al. Am J Surg Pathol 2009
6p25 gains
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Conclusion
• FISH provide additional criteria to help
diagnose histologically ambiguous
cases
• This test should be performed in
conjunction with standard
histopathologic evaluation
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CGH• Technically difficult
• Needs tumor enrichment (30-
50% tumor purity)
• Cannot be used in thin lesions
• No histologic correlations
FISH• Technically easier
• Does not need tumor
enrichment
• Can be used in thin lesions
• Allows some histologic
correlations
Vs.
• Can miss changes in
subpopulations (low sensitivity)
• Scans entire genome (improve
sensitivity)
• Positive aberrations are likely
significant (high specificity)
• Will detect focal changes (high
sensitivity)
• Evaluates only 3 loci (decrease
sensitivity)
• Higher probability of false
positive (lower specificity)– Lack of experience in counting
– Tetraploidy
– Different cutoff points
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Ambiguous lesion
Favor benign Favor malignant
FISH - FISH + FISH - FISH +
MelanomaNevus Ambiguous
(20-25% false
negative)
R/O False
Positive
•Tetraploidy
•Focal changes
Yes No
Nevus Ambiguous
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Is this final?
• Sensitivity not great
• Interpretation is labor intensive
• Technically challenging
• Expensive• Expensive
• More probes/cutoff points need to be
evaluated
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“Vidraru” Dam, Romania
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Melanoma treatment
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FDA approved therapy
• High-dose interleukin-2
• Dacarbazine
• Response rates: 7-20%• Response rates: 7-20%
• No improvement in survival
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New FDA approved drugs
• Ipilimumab (YERVOY)
– FDA approval: 3/25/2011
• Vemurafenib (ZELBORAF)• Vemurafenib (ZELBORAF)
– FDA approval: 8/17/2011
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Ipilimumab
• Blocks cytotoxic T-lymphocyte–associated
antigen 4 (CTLA4) and promoted
antitumor immune response(Q.)
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APC
MHC
Ag
TCR
T-cell
CD80/86
CD28
CD80/86
Melanoma
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APC
MHC
Ag
TCR
T-cell
CD80/86
CTLA-4
CD80/86
Melanoma
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APC
MHC
Ag
TCR
T-cell
CD80/86
CTLA-4
Melanoma
Anti
CTLA-4
AB
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Ipilimumab phase 3 trial
Ipi alone: 10.1 months
Ipi+gp100: 10 months
gp100: 6.4 months
Med survival
Hodi et al. N Eng J Med, 2010,
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Ipilimumab phase 3 trial
Ipi+dacarbazine: 11.2 months
Dacarbazine: 9.1 months
Med survival
Robert et al. N Eng J Med, 2011,
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Vemurafenib
• BRAF V600E inhibitor (Q.)
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GROWTH
FACTORS
RAS
B-RAF
CYTOPLASM
MAP Kinase pathway activation in
melanocytic lesions
40-60% of melanomas, 90% is V600E
MEK
ERK
PROLIFERATION
CYTOPLASM
NUCLEUS
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GROWTH
FACTORS
RAS
B-RAF
CYTOPLASM
MAP Kinase pathway activation in
melanocytic lesions
Vemurafenib
MEK
ERK
PROLIFERATION
CYTOPLASM
NUCLEUS
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PLX4032 (Vemurafenib)
• Phase III (BRIM3):
– 675 patients
– Vemurafenib vs Dacarbazine
– 63% reduction in risk of death
– 74% reduction in risk of death and disease
progression
Chapman PB. Et al. N Engl J Med 2011;364:2507-16.
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Vemurafenib phase 3 trial
Vemurafenib: 84%
Dacarbazine: 64%
6-month survival
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Side effects
• Rash
• Photosensitivity
• Hair loss
• Joint pain• Joint pain
• Liver problems
• Arrhythmias
• Allergic reactions
• Cutaneous SCC (26%) (Q.)
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Cobas 4800 BRAF V600E
Mutation Test• FDA approved companion diagnostic test
• Detects BRAF V600E mutation
• Real time PCR (Q.)• Real time PCR (Q.)
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Downtown, Timisoara, Romania
QUESTIONS?
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ReferencesMolecular alterations in melanoma1. Albino AP, Nanus DM, Mentle IR, Cordon-Cardo C, McNutt NS, Bressler J et al. Analysis of ras oncogenes in
malignant melanoma and precursor lesions: correlation of point mutations with differentiation phenotype.
Oncogene 1989;4:1363-74.
2. Bastian BC, LeBoit PE , Pinkel D. Mutations and copy number increase of HRAS in Spitz nevi with distinctive
histopathological features. Am J Pathol 2000;157:967-72.
3. Curtin JA, Busam K, Pinkel D , Bastian BC. Somatic activation of KIT in distinct subtypes of melanoma. J Clin
Oncol 2006;24:4340-6.
4. Curtin JA, Fridlyand J, Kageshita T, Patel HN, Busam KJ, Kutzner H et al. Distinct sets of genetic alterations in
melanoma. N Engl J Med 2005;353:2135-47.melanoma. N Engl J Med 2005;353:2135-47.
5. Davies H, Bignell GR, Cox C, Stephens P, Edkins S, Clegg S et al. Mutations of the BRAF gene in human cancer.
Nature 2002;417:949-54.
6. Haqq C, Nosrati M, Sudilovsky D, Crothers J, Khodabakhsh D, Pulliam BL et al. The gene expression signatures
of melanoma progression. Proc Natl Acad Sci U S A 2005;102:6092-7.
7. Jaeger J, Koczan D, Thiesen HJ, Ibrahim SM, Gross G, Spang R et al. Gene expression signatures for tumor
progression, tumor subtype, and tumor thickness in laser-microdissected melanoma tissues. Clin Cancer Res
2007;13:806-15.
8. Kashani-Sabet M, Rangel J, Torabian S, Nosrati M, Simko J, Jablons DM et al. A multi-marker assay to distinguish
malignant melanomas from benign nevi. Proc Natl Acad Sci U S A 2009;106:6268-72.
9. Koh SS, Opel ML, Wei JP, Yau K, Shah R, Gorre ME et al. Molecular classification of melanomas and nevi using
gene expression microarray signatures and formalin-fixed and paraffin-embedded tissue. Mod Pathol
2009;22:538-46.
10. Pollock PM, Harper UL, Hansen KS, Yudt LM, Stark M, Robbins CM et al. High frequency of BRAF mutations in
nevi. Nat Genet 2003;33:19-20.
![Page 100: Emerging Molecular Diagnostic Tests and Therapies for Melanoma](https://reader031.vdocument.in/reader031/viewer/2022020703/61fb42a12e268c58cd5c1026/html5/thumbnails/100.jpg)
ReferencesCGH/ FISH in melanoma
1. Bastian BC, LeBoit PE , Pinkel D. Mutations and copy number increase of HRAS in Spitz nevi with distinctive
histopathological features. Am J Pathol 2000;157:967-72.
2. Bastian BC, Olshen AB, LeBoit PE , Pinkel D. Classifying melanocytic tumors based on DNA copy number
changes. Am J Pathol 2003;163:1765-70.
3. Bastian BC, Wesselmann U, Pinkel D , Leboit PE. Molecular cytogenetic analysis of Spitz nevi shows clear
differences to melanoma. J Invest Dermatol 1999;113:1065-9.differences to melanoma. J Invest Dermatol 1999;113:1065-9.
4. Bastian BC, Xiong J, Frieden IJ, Williams ML, Chou P, Busam K et al. Genetic changes in neoplasms arising in
congenital melanocytic nevi: differences between nodular proliferations and melanomas. Am J Pathol
2002;161:1163-9.
5. Bauer J , Bastian BC. Distinguishing melanocytic nevi from melanoma by DNA copy number changes:
comparative genomic hybridization as a research and diagnostic tool. Dermatol Ther 2006;19:40-9.
6. Dalton SR, Gerami P, Kolaitis NA, Charzan S, Werling R, LeBoit PE et al. Use of fluorescence in situ hybridization
(FISH) to distinguish intranodal nevus from metastatic melanoma. Am J Surg Pathol 2010;34:231-7.
7. Gaiser T, Kutzner H, Palmedo G, Siegelin MD, Wiesner T, Bruckner T et al. Classifying ambiguous melanocytic
lesions with FISH and correlation with clinical long-term follow up. Mod Pathol 2010;23:413-9.
8. Gerami P, Jewell SS, Morrison LE, Blondin B, Schulz J, Ruffalo T et al. Fluorescence in situ hybridization (FISH)
as an ancillary diagnostic tool in the diagnosis of melanoma. Am J Surg Pathol 2009;33:1146-56.
9. Gerami P, Mafee M, Lurtsbarapa T, Guitart J, Haghighat Z , Newman M. Sensitivity of fluorescence in situ
hybridization for melanoma diagnosis using RREB1, MYB, Cep6, and 11q13 probes in melanoma subtypes. Arch
Dermatol 2010;146:273-8.
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ReferencesCGH/ FISH in melanoma
10. Gerami P, Wass A, Mafee M, Fang Y, Pulitzer MP , Busam KJ. Fluorescence in situ hybridization for
distinguishing nevoid melanomas from mitotically active nevi. Am J Surg Pathol 2009;33:1783-8.
11. Maize JC, Jr., McCalmont TH, Carlson JA, Busam KJ, Kutzner H , Bastian BC. Genomic analysis of blue nevi and
related dermal melanocytic proliferations. Am J Surg Pathol 2005;29:1214-20.
12. Moore SR, Persons DL, Sosman JA, Bobadilla D, Bedell V, Smith DD et al. Detection of copy number alterations
in metastatic melanoma by a DNA fluorescence in situ hybridization probe panel and array comparative genomic in metastatic melanoma by a DNA fluorescence in situ hybridization probe panel and array comparative genomic
hybridization: a southwest oncology group study (S9431). Clin Cancer Res 2008;14:2927-35.
13. Morey AL, Murali R, McCarthy SW, Mann GJ , Scolyer RA. Diagnosis of cutaneous melanocytic tumours by four-
colour fluorescence in situ hybridisation. Pathology 2009;41:383-7.
14. Newman MD, Lertsburapa T, Mirzabeigi M, Mafee M, Guitart J , Gerami P. Fluorescence in situ hybridization as a
tool for microstaging in malignant melanoma. Mod Pathol 2009;22:989-95.
15. Newman MD, Mirzabeigi M , Gerami P. Chromosomal copy number changes supporting the classification of
lentiginous junctional melanoma of the elderly as a subtype of melanoma. Mod Pathol 2009;22:1258-62.
16. Pouryazdanparast P, Newman M, Mafee M, Haghighat Z, Guitart J , Gerami P. Distinguishing epithelioid blue
nevus from blue nevus-like cutaneous melanoma metastasis using fluorescence in situ hybridization. Am J Surg
Pathol 2009;33:1396-400.
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References
Ipilimumab
1. Hodi FS, O'Day SJ, McDermott DF, Weber RW, Sosman JA, Haanen JB et al.
Improved survival with ipilimumab in patients with metastatic melanoma. N Engl J
Med 2010;363:711-23.
2. Robert C , Ghiringhelli F. What is the role of cytotoxic T lymphocyte-associated
antigen 4 blockade in patients with metastatic melanoma? Oncologist 2009;14:848-
61.
3. Robert C, Thomas L, Bondarenko I, O'Day S, M DJ, Garbe C et al. Ipilimumab plus
dacarbazine for previously untreated metastatic melanoma. N Engl J Med
2011;364:2517-26.
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ReferencesVemurafenib1. Arkenau HT, Kefford R , Long GV. Targeting BRAF for patients with melanoma. Br J
Cancer 2011;104:392-8.
2. Capper D, Preusser M, Habel A, Sahm F, Ackermann U, Schindler G et al.
Assessment of BRAF V600E mutation status by immunohistochemistry with a
mutation-specific monoclonal antibody. Acta Neuropathol 2011;122:11-9.
3. Chapman PB, Hauschild A, Robert C, Haanen JB, Ascierto P, Larkin J et al.
Improved survival with vemurafenib in melanoma with BRAF V600E mutation. N EnglImproved survival with vemurafenib in melanoma with BRAF V600E mutation. N Engl
J Med 2011;364:2507-16.
4. Flaherty KT, Puzanov I, Kim KB, Ribas A, McArthur GA, Sosman JA et al. Inhibition
of mutated, activated BRAF in metastatic melanoma. N Engl J Med 2010;363:809-19.
5. Nazarian R, Shi H, Wang Q, Kong X, Koya RC, Lee H et al. Melanomas acquire
resistance to B-RAF(V600E) inhibition by RTK or N-RAS upregulation. Nature
2010;468:973-7.
6. Ribas A, Kim KB, Schuchter LM, et al. BRIM-2: an open-label, multicenter phase II
study of vemurafenib in previousl treated patients with BRAFV600E mutationpositive
melanoma. J Clin Oncol 2011; 29:Suppl:8509. abstract.
7. Vultur A, Villanueva J , Herlyn M. Targeting BRAF in advanced melanoma: a first step
toward manageable disease. Clin Cancer Res 2011;17:1658-63.
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ReferencesOther1. Corona R, Mele A, Amini M, De Rosa G, Coppola G, Piccardi P et al. Interobserver
variability on the histopathologic diagnosis of cutaneous melanoma and other
pigmented skin lesions. J Clin Oncol 1996;14:1218-23.
2. Farmer ER, Gonin R , Hanna MP. Discordance in the histopathologic diagnosis of
melanoma and melanocytic nevi between expert pathologists. Hum Pathol
1996;27:528-31.
3. Greider CW , Blackburn EH. A telomeric sequence in the RNA of Tetrahymena 3. Greider CW , Blackburn EH. A telomeric sequence in the RNA of Tetrahymena
telomerase required for telomere repeat synthesis. Nature 1989;337:331-7.
4. Kornstein MJ , Byrne SP. The medicolegal aspect of error in pathology: a search of
jury verdicts and settlements. Arch Pathol Lab Med 2007;131:615-8.
5. Lodha S, Saggar S, Celebi JT , Silvers DN. Discordance in the histopathologic
diagnosis of difficult melanocytic neoplasms in the clinical setting. J Cutan Pathol
2008;35:349-52.
6. Lundblad V , Szostak JW. A mutant with a defect in telomere elongation leads to
senescence in yeast. Cell 1989;57:633-43.
7. McGinnis KS, Lessin SR, Elder DE, Guerry Dt, Schuchter L, Ming M et al. Pathology
review of cases presenting to a multidisciplinary pigmented lesion clinic. Arch
Dermatol 2002;138:617-21.