New Insights on Merkel Cell Carcinoma Emory Melanoma / Skin Cancer Symposium Saturday, Feb 27, 2016; Emory Conference Center 35 min talk + 10 min questions Clinic: Seattle Cancer Care Alliance Fred Hutchinson Cancer Research Center Paul Nghiem, MD, PhD Professor & Head, UW Dermatology/Medicine & Pathology George F. Odland Endowed Chair in Dermatology Research: Univ of WA Lake Union Campus UV DNA Damage/Cell cycle Merkel cell carcinoma
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New Insights on Merkel Cell Carcinoma
Emory Melanoma / Skin Cancer Symposium
Saturday, Feb 27, 2016; Emory Conference Center35 min talk + 10 min questions
Clinic:
Seattle Cancer Care Alliance
Fred Hutchinson Cancer
Research Center
Paul Nghiem, MD, PhDProfessor & Head, UW Dermatology/Medicine & Pathology
George F. Odland Endowed Chair in Dermatology
Research:
Univ of WA
Lake Union Campus
UV DNA Damage/Cell cycle
Merkel cell carcinoma
Overview
• Pathogenesis & the Merkel polyomavirus– Risk factors
– Viral oncogenesis
Overview
• Pathogenesis & the Merkel polyomavirus– Risk factors
– Viral oncogenesis
• Tumor / viral immunity– Humoral and cellular immunity
Overview
• Pathogenesis & the Merkel polyomavirus– Risk factors
– Viral oncogenesis
• Tumor / viral immunity– Humoral and cellular immunity
• Therapy– Current & future immune-based therapies
Why is MCC important?
• More lethal than melanoma– ~40% mortality (~15% for melanoma)
Why is MCC important?
• More lethal than melanoma– ~40% mortality (~15% for melanoma)
• Reported incidence increasing– Quadrupled since 1986
– Currently ~2,000 new cases/yr in USA
Why is MCC important?
• More lethal than melanoma– ~40% mortality (~15% for melanoma)
• Reported incidence increasing– Quadrupled since 1986
– Currently ~2,000 new cases/yr in USA
• Optimal therapy is unique among skin CAs– Proper tx is relatively benign, effective
What is a Merkel Cell?Friedrich Merkel, 1875 Mechanoreceptors in basal
layer of epidermis?
What is a Merkel Cell?
copyright: Paul Nghiem & Paul Quade, www.EchoMedicalMedia.com
Friedrich Merkel, 1875 Mechanoreceptors in basal
layer of epidermis?
What is a Merkel Cell?
• Essential for light touchMaricich, et al, Science, 2009
• Derived from epidermis (not neural crest)
Van Keymeulen, et al, J Cell Biol, 2009
Morrison, et al, Dev Biol, 2009
copyright: Paul Nghiem & Paul Quade, www.EchoMedicalMedia.com
Friedrich Merkel, 1875 Mechanoreceptors in basal
layer of epidermis?
Merkel cell CA: clinical appearance (easy to miss)
Clinician’s Impression at the Time of
Bx in 106 MCC pts
BENIGN 56% • Cyst/Acneiform lesion 32%
• Lipoma 6%
• Dermatofibroma 5%
MALIGNANT 36% • Non-melanoma skin CA 19%
• Lymphoma 6%
• Metastatic Carcinoma 2%
• MCC 1%
Indeterminate 8% • "Nodule" Heath, JAAD, 2008
"Perinuclear dot pattern"
of cytokeratin-20
MCC Reported Incidence is Rising
1500 new
cases/yr
in USA
in 2005(NCDB data)
Lemos & Nghiem
JID 2007
CK20
antibody
Many cases
‘missed’
1600 new
cases/yr
in USA
in 2006
(SEER data)
Albores-Saavedra
J Cut. Path 2009
Increase in Risk Factors
• Prolonged sun exposure
– 98% Caucasian; Latitude associated; sun-
exposed skin
• Immune suppression (HIV, SOTR, CLL)
• Age > 50
Paulson & Nghiem,
unpublished
Clinical appearance (easy to miss)
Clinical Presentation
Generic description: "Firm papule / nodule, often red or purple"
Adapted from Bhatia/Afanasiev, et al, Curr Oncol Rep, 2011
Virus on us all yet only
1 in 3000 will get MCC…how?
Adapted from Bhatia/Afanasiev, et al, Curr Oncol Rep, 2011
What are mutation patterns in virus-pos vs virus-neg tumors?
Virus on us all yet only
1 in 3000 will get MCC…how?
3 studies in late 2015...
genetic mutations in virus-pos & virus-neg MCC...
3 studies in late 2015...
genetic mutations in virus-pos & virus-neg MCC...
3 studies in late 2015...
genetic mutations in virus-pos & virus-neg MCC...
49 MCCs...
Goh, et al. 2015.
Oncotarget
“MCC-Low”“MCC-High”
49 MCCs...
Goh, et al. 2015.
Oncotarget
“MCC-Low”“MCC-High”
49 MCCs...
Goh, et al. 2015.
Oncotarget
“MCC-Low”“MCC-High”
Goh, et al. 2015.
Oncotarget
Viral status
greatly affects
mutation
frequency
Goh, et al. 2015.
Oncotarget
Viral status
greatly affects
mutation
frequency
100-fold
difference
between
MCC-Lo &
MCC-Hi
Goh, et al. 2015. Oncotarget
Virus-negative MCCs ‘neoantigens’
PDL1-pos (‘immune visible’) tumors
have more mutations
Wong, et al. 2015.
Cancer Res.
PDL1-pos (‘immune visible’) tumors
have more mutations
Wong, et al. 2015.
Cancer Res.
Virus-neg tumors
may be
immunogenic...
We will return to
this!
Humoral immunity: a powerful biomarker
Kelly Paulson, MD, PhD Will Simonson,MD, PhDMark Wener, MDDenise Galloway, PhD Jody Carter, PhD
Antibodies to T-Ag (not capsid) fall after tx
Antibodies to T-Ag (not capsid) fall after txCapsid antibodies:>60% of us have them!
T-Ag antibodies:<1% of 530 population controls~50% of newly diagnosed MCCs
Antibodies to T-Ag (not capsid) fall after txCapsid antibodies:>60% of us have them!
T-Ag antibodies:<1% of 530 population controls~50% of newly diagnosed MCCs
Antibodies to T antigen reflect disease burden
Paulson, Carter, et al, CA Research, 2010
“This could be clinically useful...”
Detecting MCC
recurrences using
serology
Paulson, et al, submitted
PPV = 66%
NPV = 97%
Oncoprotein Ab Positive (n = 114)
Oncoprotein Ab Negative (n = 105)
Competing-risks Regression
Controlling for: Age, Stage, and Sex
p = 0.037
Hazard Ratio = 0.46
95% CI = 0.224 – 0.955
Time after Diagnosis (Days)
MC
C s
pe
cif
ic S
urv
iva
l (%
)MCC Specific Survival by Oncoprotein Ab Titer
590 1 2 3 4 5
0
25
50
75
100
Paulson, et al, submitted
Oncoprotein Ab Positive (n = 114)
Oncoprotein Ab Negative (n = 105)
Competing-risks Regression
Controlling for: Age, Stage, and Sex
p = 0.037
Hazard Ratio = 0.46
95% CI = 0.224 – 0.955
Time after Diagnosis (Days)
MC
C s
pe
cif
ic S
urv
iva
l (%
)MCC Specific Survival by Oncoprotein Ab Titer
600 1 2 3 4 5
0
25
50
75
100
Sero-neg patients:
follow with scans
Paulson, et al, submitted
Assay ‘live’ (available) as of January 2014...Run by UW Lab Medicine (50 ul serum)
Cost $200 (very modest vs CT scan)Helps both virus-pos and virus-neg patients...
Antibodies are useful for tracking MCC...
What are T cells good for?
Antibodies are useful for tracking MCC...
What are T cells good for?
Survival!!
If CD8+ T cells are moderate or high within tumor,survival is 100% (n=146 patients + n=150 patients)
killer CD8 T cell
MCC cell
expressing MCPyV protein
How do CD8 T
cells recognize
their target?
Finding MCPyV-specific T cells
x
xx
x
MCPyVproteome
map
MCPyV-Antigen source
Jayasri Iyer
David Koelle
Finding MCPyV-specific T cells
x
xx
x
MCPyVproteome
map
MCPyV-Antigen source
95 peptides: covering 389 AA
Persistently expressed in MCCs
Jayasri Iyer
David Koelle
T cell source
Finding MCPyV-specific T cells
x
xx
x
MCPyVproteome
map
MCPyV-Antigen source
95 peptides: covering 389 AA
Blood “TIL”Persistently expressed in MCCs
Jayasri Iyer
David Koelle
T cell source
Antigen presenting cells
Finding MCPyV-specific T cells
x
xx
x
MCPyVproteome
map
MCPyV-Antigen source
95 peptides: covering 389 AA
Blood “TIL”
Assay: T cells produce IFN-γ if recognize peptide
Persistently expressed in MCCs
Jayasri Iyer
David Koelle
95 peptides: covering 389 AA
x
xx
x
MCPyVproteome
map
MCPyV
Persistently expressed in MCCs
First 26 MCPyV epitopes identified from blood and MCC tumors
epitopesdiscovered
‘Tools’ for MVPyV-specific T cell study
Iyer/Afanasiev et al, CCR, 2011; Afanasiev et al, submitted
Iyer, et al, Clin Ca Res, 2011
‘Tools’ for MVPyV-specific T cell study
Iyer/Afanasiev et al, CCR, 2011; Afanasiev et al, submitted
Control subjects
(n=10)
% s
ub
jects
with
te
t+T
ce
lls
MCC patients
(n=9)
0%
Iyer, et al, Clin Ca Res, 2011
Jim Dowdalls/Photo Researchers, Inc
Why do virus-positive tumors grow if patients have virus-specific T cells??
Virus infected cell
CD8T cell
Are virus-specific T cells dysfunctional?
Acute antigen
exposure
Activated cells, characterized by expression of:
CD28 Co-stimulatory receptor (ligand: B7); required for T cell activation
CD69 Earliest inducible cell surface glycoprotein during T cell activation; plays a role in T cell proliferation
CD137 (4-1BB)
Member of TNF-receptor family; induced by T cell activation; important in T cell proliferation, cytokine secretion and cytotoxicity
CD38 Cyclic ADP ribose hydrolase; marker of T cell activation; functions in cell adhesion, signal transduction and calcium signaling
HLA-DR MHC class-II surface receptor that is upregulated with T cell activation
T cell surface receptor phenotype
reveals functional profile
Adapted from Afanasiev, Nghiem, MCC book chapter
Acute antigen
exposure
Activated cells, characterized by expression of:
CD28 Co-stimulatory receptor (ligand: B7); required for T cell activation
CD69 Earliest inducible cell surface glycoprotein during T cell activation; plays a role in T cell proliferation
CD137 (4-1BB)
Member of TNF-receptor family; induced by T cell activation; important in T cell proliferation, cytokine secretion and cytotoxicity
CD38 Cyclic ADP ribose hydrolase; marker of T cell activation; functions in cell adhesion, signal transduction and calcium signaling
HLA-DR MHC class-II surface receptor that is upregulated with T cell activation
T cell surface receptor phenotype
reveals functional profile
Adapted from Afanasiev, Nghiem, MCC book chapter
Chronic antigen
exposure
Acute antigen
exposure
Activated cells, characterized by expression of:
CD28 Co-stimulatory receptor (ligand: B7); required for T cell activation
CD69 Earliest inducible cell surface glycoprotein during T cell activation; plays a role in T cell proliferation
CD137 (4-1BB)
Member of TNF-receptor family; induced by T cell activation; important in T cell proliferation, cytokine secretion and cytotoxicity
CD38 Cyclic ADP ribose hydrolase; marker of T cell activation; functions in cell adhesion, signal transduction and calcium signaling
HLA-DR MHC class-II surface receptor that is upregulated with T cell activation
Exhausted T cells, characterized by prolonged expression of:
PD-1 Programmed death-1; inhibitory T cell receptor (ligands: PD-L1 (B7-H1), PD-L2 (B7-DC)); reduces T cell proliferation and effector functions
CTLA-4 (CD152)
Cytotoxic T-Lymphocyte Antigen 4; inhibitory receptor (ligand:B7); effectively competes for ligands with CD28 (which has lower avidity than CTLA-4), preventing T cell activation
Tim-3 T cell Immunoglobulin Mucin-3; inhibitory T cell receptor (ligand: galactin-9); leads to decrease in effector T cell function
T
cell
T cell surface receptor phenotype
reveals functional profile
Adapted from Afanasiev, Nghiem, MCC book chapter
Chronic antigen
exposure
Acute antigen
exposure
Activated cells, characterized by expression of:
CD28 Co-stimulatory receptor (ligand: B7); required for T cell activation
CD69 Earliest inducible cell surface glycoprotein during T cell activation; plays a role in T cell proliferation
CD137 (4-1BB)
Member of TNF-receptor family; induced by T cell activation; important in T cell proliferation, cytokine secretion and cytotoxicity
CD38 Cyclic ADP ribose hydrolase; marker of T cell activation; functions in cell adhesion, signal transduction and calcium signaling
HLA-DR MHC class-II surface receptor that is upregulated with T cell activation
Exhausted T cells, characterized by prolonged expression of:
PD-1 Programmed death-1; inhibitory T cell receptor (ligands: PD-L1 (B7-H1), PD-L2 (B7-DC)); reduces T cell proliferation and effector functions
CTLA-4 (CD152)
Cytotoxic T-Lymphocyte Antigen 4; inhibitory receptor (ligand:B7); effectively competes for ligands with CD28 (which has lower avidity than CTLA-4), preventing T cell activation
Tim-3 T cell Immunoglobulin Mucin-3; inhibitory T cell receptor (ligand: galactin-9); leads to decrease in effector T cell function
Recently activated T cells, characterized by expression of:
Combination of activation and inhibition markers via appropriate immunoregulatory feedback mechanisms
T
cell
T cell surface receptor phenotype
reveals functional profile
Adapted from Afanasiev, Nghiem, MCC book chapter
Chronic antigen
exposure
Acute antigen
exposure
Activated cells, characterized by expression of:
CD28 Co-stimulatory receptor (ligand: B7); required for T cell activation
CD69 Earliest inducible cell surface glycoprotein during T cell activation; plays a role in T cell proliferation
CD137 (4-1BB)
Member of TNF-receptor family; induced by T cell activation; important in T cell proliferation, cytokine secretion and cytotoxicity
CD38 Cyclic ADP ribose hydrolase; marker of T cell activation; functions in cell adhesion, signal transduction and calcium signaling
HLA-DR MHC class-II surface receptor that is upregulated with T cell activation
Exhausted T cells, characterized by prolonged expression of:
PD-1 Programmed death-1; inhibitory T cell receptor (ligands: PD-L1 (B7-H1), PD-L2 (B7-DC)); reduces T cell proliferation and effector functions
CTLA-4 (CD152)
Cytotoxic T-Lymphocyte Antigen 4; inhibitory receptor (ligand:B7); effectively competes for ligands with CD28 (which has lower avidity than CTLA-4), preventing T cell activation
Tim-3 T cell Immunoglobulin Mucin-3; inhibitory T cell receptor (ligand: galactin-9); leads to decrease in effector T cell function
Recently activated T cells, characterized by expression of:
Combination of activation and inhibition markers via appropriate immunoregulatory feedback mechanisms
T
cell
T cell surface receptor phenotype
reveals functional profile
Adapted from Afanasiev, Nghiem, MCC book chapter
Are the ‘brakes’ ON in MCC-specific T cells? (compared to other viral responses)
PD-1 Tim-3
Examined ‘exhausted’ PD1+/Tim-3+ cells
Are the ‘brakes’ ON in MCC-specific T cells? (compared to other viral responses)
Afanasiev et al, Clin Ca Research 2013
PD
1+T
im3
+ (%
po
siti
ve c
ells
)PD-1 Tim-3
(Data from 4-7 MCC patients)
Examined ‘exhausted’ PD1+/Tim-3+ cells
Afanasiev et al, Clin Ca Research 2013
PD
1+T
im3
+ (%
po
siti
ve c
ells
)PD-1 Tim-3
(Data from 4-7 MCC patients)
Examined ‘exhausted’ PD1+/Tim-3+ cells
Are the ‘brakes’ ON in MCC-specific T cells? (compared to other viral responses)
Afanasiev et al, Clin Ca Research 2013
PD
1+T
im3
+ (%
po
siti
ve c
ells
)PD-1 Tim-3
(Data from 4-7 MCC patients)
Examined ‘exhausted’ PD1+/Tim-3+ cells
PD-1 & PDL-1 trials
are beginning...
Are the ‘brakes’ ON in MCC-specific T cells? (compared to other viral responses)
55 yo woman; primary lesion on chest x 1 year (had metastases at dx)
Failed chemotherapy (Carbo + etoposide)
Large liver mets: 2 x 11 cm
MCPyV-negative
After 4 weeks (2 doses) of PDL1
Activity of PD-1 blockade with pembrolizumab as first systemic therapy in patients with advanced
Merkel cell carcinoma
Paul Nghiem1,2, Shailender Bhatia1,2, Adil Daud3, Philip Friedlander4, Harriet Kluger5, Holbrook Kohrt6, Ragini Kudchadkar7, Evan Lipson8, Lisa Lundgren2,
Kim Margolin6, Sunil Reddy6, Erica Shantha1, William Sharfman8, Elad Sharon9, John Thompson1,2, Suzanne Topalian8, Mac Cheever1,2
1) University of Washington 2) Fred Hutchinson Cancer Research Center 3) Univ of California San Francisco 4) Mt Sinai Medical Center 5) Yale University
6) Stanford University 7) Emory University 8) Johns Hopkins University