Immunotherapy :

A New Approach of Cancer Treatment

AMI ASHARIATI
Division of Hematology and Medical Oncology
Department of Internal Medicine
Airlangga University School of Medicine – Dr. Soetomo Hospital
Surabaya
 Basic knowledge of cancer

• What is Cancer?
• How does cancer occur?
• How many types of cancers?
• Immunosurveillance
• Immunotherapy (Current therapeutic strategies) for
cancers
 Definitions of Cancer

• Cancer consists of single clones or several

clones of cells that are capable of independent
growth in the host
• Cancer cells arise from host cells via neoplastic
transformation or carcinogenesis.
• Physical, chemical and biological agents may cause
cancer  carcinogens
 Carcinogens
• Radiation: Ultraviolet light, sunshine; X-rays,
radioactive elements induce DNA damage and
chromosome brakes.
• Chemical: smoke and tar, countless chemicals
that damage DNA (mutagens).
• Oncogenic viruses: insert DNA or cDNA copies
of viral oncogens into the genome of host target
cells (EBV)
• Hereditary: certain oncogenes are inheritable
 Neoplastic Transformation
-- Carcinogenesis
• Activate growth regulatory genes:
-Growth factor receptors (erbA, -B, fims, neu);
-molecules of signal transduction (src, abl, ras);
-transcription factors (jun, fos ,myc)
• Genes that inhibit growth:
-p53 controls DNA repair and cell proliferation;
-Rb – suppressor oncogenes
• Genes that regulate apoptosis: bcl-2, Bax, Bid.
 Classification of cancer
• Carcinoma : -epithelial origin involving the skin,
-mucous membranes,
-epithelial cells in glands
• Sarcoma : -cancer of connective tissue, i.e.
liposarcoma, fibrosarcoma
• Lymphoma :- T-cell, B-cell, Hodgkin’s,
- Burkitt’s lymphomas
• Leukemia : disseminated tumors - lymphoid,
myeloid, acute and chronic
 The hallmarks of cancer

• Growth self-sufficiency
• peran sistem imun spesifik maupun non spesifik
• Evade
dalam apoptosis
mencegah pertumbuhan tumor spontan dan
• Ignore anti-proliferative
• bagaimana signals peran
memodulasinya, memegang
penting di kemudian
• Limitless hari dalam:
replication potential
- meningkatkan surveillance terhadap tumor,
• Sustained angiogenesis
- menginduksi resistensi terhadap sisa sel ganas
• Invade tissues
- menghambat perkembangan tumor (relaps),
•- Escape immune
menentukan surveillance
jenis pengobatan.
 Cancer Immuno-surveilance

• The hypothesis was first proposed by Ehrlich in 1909,

and modified by Thomas and Burnet in 1957.
• Immunosurveilance: the immunological resistance of
the host against the development of cancer.
• Now referred to “cancer immunoediting”
encompassing 3 phases: elimination, equilibrium and
escape
 Topic

• How does the immune system eliminate cancer cells?

• How do cancer cells escape from immunosurveilance?
• How can we help to win the battle between immune
system and cancers? – Cancer immunotherapy
• Examples
 macrophage

• Gambar 1. Mekanisme imunitas alamiah/innate/non-spesifik memulai perlawanan terhadap

infeksi. Imunitas didapat/acquired/spesifik berkembang kemudian dan mengandung limfosit.
Aktifitas respon imun alami dan didapat bervariasi pada berbagai bentuk infeksi, termasuk
terhadap tumor.
Innate:
Macrop/NK/NKT/ϒϬ T cells
Adaptive:CD4/CD8+ T cells
IFN-ϒ/TRAIL/FasL
/perforin
TA-dependent Immune response TA-independent Immune response
TA-dependent Immune response
 Cellular Immune Response

Gambar 3. Sel T CD8+ berespon terhadap tumor dapat diinduksi oleh pengenalan
silang (cross-priming), di mana sel-sel tumor atau antigen tumor diambil, diproses, dan
dipresentasikan kepada sel T oleh sel-sel yang mempresentasikan antigen (antigen-
presenting cells=APCs). Pada beberapa kasus, ko-stimulator B7 yang diekspresikan oleh
APCs memungkinkan sinyal kedua kepada sel T CD8+. APC juga menstimulasi sel T-
helper CD4+ yang memungkinkan sinyal kedua untuk limfosit T-sitolitik (CTL). CTL
membunuh sel-sel tumor tanpa memerlukan ko-stimulator atau sel T-helper.
 The Immune System Can Fight Tumors Via
A Variety of Functionally Specialized Cells1

Dendritic cells
T cells
Myeloid- Lymphoid-
derived cells derived cells
Hematopoietic
Present antigens Eradicate
derived from Stem cell pathogens and
pathogens and nascent tumor cells
nascent tumor cells to B cells through their
immune cells, response to
including T cells, B antigens that are
cells, and not expressed in
Macrophages
NK cells normal tissue
NK cells

NK = natural killer.
1. Norvell A. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:11–24.
 Some Tumor Cells Express Multiple Antigens
That Are Not Expressed by Normal Cells1
Tumor cells release differentially
Normal cells release molecules that are
expressed antigens that cause them to
captured by antigen-presenting cells, but
be recognized as foreign entities and
they don’t elicit an immune response.
therefore elicit an immune response.

NORMAL TUMOR
CELL CELL

1. Finn OJ. N Engl J Med. 2008;358:2704–2715.

 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

TUMOR

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
2
0
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

Activated
T cell

3. Activated T cells migrate back to the

tumor.

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

Activated
4. T cells kill tumor cells through T cell
the release of lytic enzymes or
induction of apoptosis.

3. Activated T cells migrate back to the

tumor.

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
2
2
 T-Cell Activity Is Regulated By Immune
Checkpoints to Limit Autoimmunity1
Dendritic cell

Inactivated

-
T cell

-
TUMOR
Checkpoints

Checkpoints
Checkpoints
Inactivated LYMPH
T cell NODE

Immune checkpoints, such as

CTLA-4, PD-1, LAG-3, and TIM-3
function at different phases in Activated
T cell
the immune response to
regulate the duration and
level of the T-cell PD-1 is a negative co-stimulatory
response.
receptor expressed primarily on
activated T cells
CTLA-4 = cytotoxic T-lymphocyte antigen 4; PD-1 = programmed cell death protein 1; LAG-3 = lymphocyte activation gene 3; 1
TIM-3 = T-cell immunoglobulin and mucin protein 3.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–264.
2
3
 Cytokines
• Regulating the innate immune system: NK cells, macrophages,
and neutrophils; and the adaptive immune system: T and B cells
• IFN-α : upregulating MHC class I, tumor antigens, and adhesion
molecules; promoting activity of B and T cells, macrophages,
and dendritic cells.
• IL-2 : T cell growth factor that binds to a specific tripartite
receptor on T cells.
• IL-12 : promoting NK and T cell activity, and a growth factor
for B cells
• GM-CSF (Granulocyte-monocyte colony stimulating factor) :
reconstituting antigen-presenting cells.
(Innate and Adaptive immune)
 Topic
• How does the immune system eliminate cancer cells?
• How do cancer cells escape from immunosurveilance?
• How can we help to win the battle between immune
system and cancers? – Cancer immunotherapy
• Examples
 How cancer cells avoid
Immunosurveillance
1. Weapons from the tumors
2. Defects of immune system
 Mechanisms of cancer escape from
Immunosurveilance
1.Weapons from the tumors
1. Altering Their Characteristics :
- Loss/downregulation of MHC class I
- Down-regulation, mutation, or loss of tumor antigens
- Loss of costimulation
2. Suppressing the Immune Response :
- ineffective signals to CTL
- Alteration in cell death receptor signaling
- Immunosuppressive cytokine
3. Outpacing the Immune Response: Tumour cells can simply
proliferate so quickly that the immune response is not fast
enough to keep their growth in check
mutation, or loss of tumor antigens

Loss/downregulation of MHC class

Immunosuppressive cytokines
 (1) Loss/down-regulation of HLA class I
(MHC I)
• Total loss: β2 microglobulin mutation, alteration in
antigen processing machinery
• Haplotype loss: LOH in chromosome 6
• HLA allelic loss: mutations of HLA class I genes
 (2) Down-regulation, mutation or loss of
tumor antigens

• Tumor antigens (TA)

• Tumor associated antigen (TAA)

• Complete loss
• Down-modulation
 Tumor Antigens
• Altered self : * K-ras mutation,
* products of normally unexpressed genes
(MAGE, BAGE, GAGE),
* proteins of alternative reading frame of :
- post-translational modification,
- different orders of glycosylation (mucin-CA125, MUC1).
• Viral antigens : EBV-A, E-6,E-7, papilloma virus antigens of
cervical carcinomas.
• Oncofetal antigens: alpha-fetoprotein (AFP), CEA
• Autoantigens : overexpression
- c-myc in lymphomas, leukemias
- HER-2/neu epidermal growth factor receptor (BC)
 (3) Immunosuppressive Cytokines

• IL-10 inhibits APC and IL-12 production

• TGF-beta induces overproduction of IL-10
• VEGF (vascular endothelial growth factor) : avoid
immune recognition, inhibit the effector function,
prevent T cell activation, cytokine production.
 Nature of TGF-β

PSR: Phosphatidyl Serine Reseptor

 (4) Loss of costimulation

Costimulatory molecules
• B7.1 (recognized:
B7.1(CD80) B7 family
B7.2(CD86)
• CD40 L
• CD27, CD30
• 4-1BB
• OX40
• ICAM-1
CD 80 CD 86
? ?

Co-stimulation Loss of costimulation

The initiation of T cell responses requires
two distinct signals

PD-1 is a negative co-stimulatory

receptor expressed primarily on
activated T cells1

The Role of signal s1 and 2 in T- cell activation

 (5) Alterations in cell death receptor
signalling
• Defects in Fas/FasL signaling system: resistance to
apoptosis
• Apoptosis resistance : overexpression of Bcl-2
(Hodgkin Lymphoma)
(6) Induction of Immunosuppressive cells

• CD4+CD25+ T cells (constitute 5-10% of CD4+ T cells):

immunological tolerance to self-antigens, inhibition of T cell
proliferation

• Gr1+CD11b+ myeloid cells :

-- Expressing the granulocyte-monocyte markers Gr1+CD11b+,
accumulate in spleens, lymph nodes and blood of tumor
bearing mice.
-- Inhibiting antibody production, CTL generation, T cell
function, lymphocytic proliferation, CD3 ζ chain expression.
inhibitor
(7) Production of other suppressive factors

• IDO (Indoleamine 2, 3-dioxygenase):

expressed in most human tumour tissues
and splenic DC subsets, leading to blockage of
proliferation of T cells
• ganglioside (sialic acid containing glycosphingolipids)
• Prostaglandins
 Immunosuppressive regulators in
tumor microenvironment

PD-L 1 PD-1

(MDSC=Myeloid Derived supressor Cell; TAM=tumor Associated Macrophage) C.H. Kapadia et al. / Journal of Controlled Release xxx (2015) xxx–xxx
 How cancer cells avoid
Immunosurveillance

1. Weapons from the tumors

2. Defects of immune system
2. Defects of immune system

• Immune defects in T cells

• Malfunction of dendritic cell system
 Immune defects in T cells in cancer

• Alterations in the expression of signal transduction

molecules in immune cells.
• Loss of CD 3ζ chain. TCR-CD3 complex. It functions
as a single transducer upon antigen binding.
• Receptor-mediated apoptosis of T cells contributes
to T cell dysfunction.
Malfunction of dendritic cell system

– Tumor-mediated inhibition of DC generation,

differentiation and maturation
– Functional impairment of DCs: lack of expression of
costimulatory molecules
– Induction of DC apoptosis by tumors
Tumor-Immune System Crosstalk
 Topic
• How does the immune system eliminate cancer
cells?
• How do cancer cells escape from Immunosurveilance?
• How can we help to win the battle between immune
system and cancers? – Cancer immunotherapy
• Examples
Cancer Immunotherapy
Main Mechanisms of Immunotherapy

• Stimulating the antitumor response, either by

increasing the number of effector cells or by
producing soluble mediators.
• Decreasing suppressor mechanisms.
• Altering tumor cells to increase their
immunogenicity and make them more susceptible
to immunologic defences
• Improving tolerance to cytotoxic drugs or
radiotherapy, such as stimulating bone marrow
function with GM-CSF.
 History:
• 1892 - WB Coley observed tumour regression
after bacterial infections
• BCG vaccine to treat bladder carcinoma
• 1970-80’s – cytokines
– includes interferons, interleukins and tumor
necrosis factor (TNF)
- limited success
 What Have We Learned About the Role of the
Immune System in Oncology?
Burnet and Thomas:
The theory of
Coley reports cases Reports of tumor
immunosurveillance2:
of tumor regression regression in patients
The immune system
following administered LAK with Regulators of T-cell The immune
patrols the body to
inoculation with IL-2.4 activity, spurring response remains
detect and destroy
erysipelas research into the role of an active focus of
nascent tumor cells.
infection.1 immune checkpoints in cancer research.9
cancer are elucidated.6

1890s 1909 Late 1950s 1980s 1985 1990s 1995 2000s Present

Paul Ehrlich proposed a

role for the immune
system against cancer.2
Beginning in the 1980s,
immunosuppressed HIV
patients were shown to
be at increased risk of
certain cancers.3
Tumors are found to Increased
express antigens tumorigenesis in
that can elicit a T immunodeficient
cell–mediated mice lacking T-, B-
immune response.5 and NKT cells.7,8

HIV = human immunodeficiency virus; LAK = lymphokine-activated killer; IL-2 = interleukin-2; NKT = natural killer T.
1. Coley WB. Am J Med Sci. 1893;105:487–511. 2. Ichim CV. J Transl Med. 20058;3:8. 3. Levine AM et al. Curr Probl Cancer. 1987;11:209–55. 4.
Rosenberg SA et al. N Engl J Med. 1985;313:1485–1492. 5. van der Bruggen P et al. Science. 1991;254:1643–1647. 6. Tivol EA. et al. Immunity.
1995;3:541–547. 7. Vesely MD et al. Annu Rev Immunol. 2011;29:235–271. 8. Shankaran V. et al. Nature. 2001;410:1107–1111. 9. Drake CG et al.
Nat. Rev. Clin. Oncol. 2014;11: 24–37.
5
6
 Current Immunotherapeutic
strategies in clinic or clinical trials

• Antibody Therapy (Unconjugated Mo-Ab; anti-idiotype Mo-Ab)

• Cytokine Therapy ( Interferon; IL-2 )
• Cellular Adoptive Therapy
- immune checkpoint inhibition (PD-1; PD-L1; CTLA-4 inh)
- genetic Engineering approach ( CAR-T cell –
Chimeric Antigen receptor T cell )
• Vaccination (DC vaccine ?)
• Combinational therapy
Immunotherapeutic strategies for cancer

C.H. Kapadia et al. / Journal of Controlled Release xxx (2015)

xxx–xxx
 Examples of FDA-approved cancer
immunotherapeutic agents
Modality Principle
Hematopoietic stem cell transplantion
(e.g., leukemia and myeloma) 1. Reset the immune system
2. Allo-antigen response
(graft versus tumor
effect)
Antibody (e.g., retuximab, trastuzumab) 1. Eliminate cancer cells
2. Block key signaling
pathways
Cytokines (e.g., type I interferon, interleukin-2) Boost both innate and adaptive
immunity
Dendritic cells (e.g., Sip-T for prostate cancer) Enhance tumor-specific T cell
priming
T cell checkpoint blockade (e.g., Ipilimumab
for melanoma) Block/reverse immune
tolerance

Microbes (e.g., BCG for the transitional

BREAST Ca,
NSCLC, NHL
Anti PD-1
Blocking antibodies to CTLA-4
VACCINE
 The Immune System Can Fight Tumors Via
A Variety of Functionally Specialized Cells1

Dendritic cells
T cells
Myeloid- Lymphoid-
derived cells derived cells
Hematopoietic
Present antigens Eradicate
derived from Stem cell pathogens and
pathogens and nascent tumor cells
nascent tumor cells to B cells through their
immune response to
cells, including T antigens that are
cells, B cells, and not expressed in
Macrophages
NK cells normal tissue
NK cells

NK = natural killer.
1. Norvell A. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:11–24.
 Some Tumor Cells Express Multiple Antigens
That Are Not Expressed by Normal Cells1
Tumor cells release differentially
Normal cells release molecules that are
expressed antigens that cause them to
captured by antigen-presenting cells, but
be recognized as foreign entities and
they don’t elicit an immune response.
therefore elicit an immune response.

NORMAL TUMOR
CELL CELL

1. Finn OJ. N Engl J Med. 2008;358:2704–2715.

 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

TUMOR

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
7
2
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

Activated
T cell

3. Activated T cells migrate back to the

tumor.

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

Activated
4. T cells kill tumor cells through T cell
the release of lytic enzymes or
induction of apoptosis.

3. Activated T cells migrate back to the

tumor.

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
7
4
 T-Cell Activity Is Regulated By Immune
Checkpoints to Limit Autoimmunity1
Dendritic cell

Inactivated

-
T cell

-
TUMOR
Checkpoints

Checkpoints
Checkpoints
Inactivated LYMPH
T cell NODE

Immune checkpoints, such as

CTLA-4, PD-1, LAG-3, and TIM-3
function at different phases in Activated
T cell
the immune response to
regulate the duration and
level of the T-cell
response.
CTLA-4 = cytotoxic T-lymphocyte antigen 4; PD-1 = programmed cell death protein 1; LAG-3 = lymphocyte activation gene 3;
TIM-3 = T-cell immunoglobulin and mucin protein 3.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–264.
7
5
 Exploiting the PD-1 Immune Checkpoint Pathway

Priming Phase of Effector Phase

Activation

Dendritic cell Naïve T cell Activated T cell Tumor cell

• Emerging research has

PD-1 identified PD-1 as an
immune checkpoint
pathway that tumor cells
may exploit to evade
Antigen
immune surveillance
• Tumor cells may block
immune responses via the
TCR MHC PD-1 immune checkpoint
pathway by expressing the
dual PD-1 ligands, PD-L1
and PD-L2
PD-L1

PD-1
Adapted with permission from Pardoll DM.1 PD-L2

PD-1 = programmed cell death protein 1; TCR = T-cell receptor; MHC = major histocompatibility complex; PD-L1 = programmed
cell death ligand 1; PD-L2 = programmed cell death ligand 2.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–64.
 Exploiting the PD-1 Immune Checkpoint
Pathway
Priming Phase of Effector Phase
Activation
Dendritic cell Naïve T cell Inactivated T cell Tumor cell

-
• PD-1 is upregulated on
PD-1 activated T cells during
the effector phase of the
immune response
Antigen • PD-L1 and PD-L2
engage the PD-1
TCR MHC receptor on T cells to
downregulate T-cell

-
activity in the effector
PD-L1 phase
PD-1

PD-L2

T cell Tumor cells

Inactivation evade destruction
Adapted with permission from Pardoll DM.1
PD-1 = programmed cell death protein 1; PD-L1 = programmed cell death ligand 1; PD-L2 = programmed cell death ligand 2.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–264.
 PD-1 Pathway and Pembrolizumab
• PD-1 is a negative co-stimulatory
receptor expressed primarily on
activated T cells1
• Binding of PD-1 to its ligands
PD-L1 and PD-L2 inhibits effector
T-cell function1
• PD-L1 expression on tumor cells and
macrophages suppresses immune
surveillance and permits neoplastic
growth2
• Pembrolizumab is a humanized
monoclonal IgG4 antibody
– Binds to PD-1 with high affinity
– Prevents PD-1 from binding to PD-L1
and PD-L2
– Robust antitumor activity and
manageable in multiple advanced
malignanciesa

aFDA-approved for the treatment of patients with unresectable or metastatic melanoma and disease progression following ipilimumab and, if BRAF V600 mutation
positive, a BRAF inhibitor.
1. Keir ME et al. Annu Rev Immunol. 2008;26:677-704; 2. Pardoll DM. Nat Rev Cancer. 2012;12:252-64.
Anti PD-1
 PD-L1 and PD-L2 Can Be Expressed on
Some Tumor Cells
 PD-L1 is expressed on some tumor cells and may
downregulate tumor-specific T-cell activity by binding to
PD-1 in the tumor microenvironment1–5
 – In some tumors, high PD-L1 expression based on immunohistochemistry
has been associated with a poor prognosis6–8

 PD-L2 may also play a role in helping tumor cells evade

the immune response8–11
 The role of immunology in cancer, including PD-1 and
its dual ligands PD-L1 and PD-L2, is a significant
focus in oncology research
PD-L1 = programmed cell death ligand 1; PD-L2 = programmed cell death ligand 2; PD-1 = programmed cell death protein 1.
1. Hino R et al. Cancer. 2010;116:1757–1766; 2. Wintterle S et al. Cancer Res. 2003;63:7462–7467; 3. Konishi J et al. Clin Cancer Res. 2004;10:5094–
5100; 4. Hamanishi J et al. Proc Natl Acad Sci U S A. 2007;104:3360–3365; 5. Inman BA et al. Cancer. 2007;109:1499–1505; 6. Loos M et al. Ann Thorac
Surg. 2011;91:1025–1031; 7. Thompson RH et al. Cancer Res. 2006;66:3381–3385; 8. Nomi T et al. Clin Cancer Res. 2007;13:2151–2157; 9. Ghebeh H
et al. Neoplasia. 2006;8:190–198; 10. Chapon M et al. J Invest Dermatol. 2011;131:1300–1307; 11. Karim R et al. Clin Cancer Res. 2009;15:6341–6347
 PD-L1: Investigational Biomarker
for anti–PD-(L)1 Immunotherapy
PD-L1 Expression by NSCLC
• Overexpressed in NSCLC Correlates With Poor Therapy Outcome5
and has potential to be used as a
biomarker1-3 PD-L1 (+) PD-L1 (–)
1.0
• Associated with poor prognosis1-
3
0.8
• Associated with higher response
rates to anti–PD-(L)1 therapy1

Overall Survival
0.6

• PD-L1 IHC remains the only 0.4

near-term companion diagnostic
option for drugs targeting in the 0.2

PD-1 pathway4
0.0

0 10 20 30 40 50
Month After Surgery
IHC, immunohistochemistry; PD-1, programmed cell death 1; PD-L1, programmed cell death ligand 1.

1. Gandhi L et al. Presented at: AACR 2014; April 6, 2014. Abstract CT105. 2. Zhang Y et al. Onco Targets Ther. 2014:7(12):567–573.
3. Chen Y-b et al. Tumori. 2012;98(6):751-755. 4. Sznol M, Chen L. Clin Cancer Res. 2013;19(5):1021-1034. 5. Mu C-Y et al. Med Oncol. 2011;28(3):682-688.
Anti T lymphocyte : CTLA4- Ig recombinant protein

MHC class II T cell activation :  Antigen generates ( signal 1)

TCR
 CD28 co-stimulation provides (signal 2)
T cell

APC T Cell Effect

Antigen
Signal 1
CD40 CD40-L +
PD-1L PD-1 -
ICOS-L ICOS +
CD80/86 CTLA4 -
Activated
T cell MHC TCR +
CD80/86 CD28 +
CD80/86CD28
PD-2L ? ?
Signal 2 B7-H3 ? ?

DC = dendritic cell
Moreland (2004).www.medscape.com
Ipilimumab
 Summary (1) :
Main defences of the tumors against immunity
1) Alteration of MHC class I and tumor antigen expression
2) Dysregulated expression of adhesion / costimulatory
molecules by tumor and/or antigen-presenting cells
3) Changes in T-cell signal transduction molecules, i.e. cell
death receptor signalling
4) Induction of immune suppressive cytokines
5) Induction of immunosuppressive cells
6) Secretion of suppressive factors

Immunotherapy may be the next great hope for

cancer treatment
 SUMMARY (2) :

•Immunotherapy may be the next great hope for

cancer treatment.
• While antibodies, cytokines, and vaccines have
individually shown some promise, it is likely that
the best strategy to combat cancer will be to
attack on all fronts.
• The effect of immunotherapy in combination with
traditional cancer therapies is another avenue.
 TERIMA KASIH

SURABAYA
 TERIMA KASIH

SURABAYA
 The Immune System Can Fight Tumors Via
A Variety of Functionally Specialized Cells1

Dendritic cells
T cells
Myeloid- Lymphoid-
derived cells derived cells
Hematopoietic
Present antigens Eradicate
derived from Stem cell pathogens and
pathogens and nascent tumor cells
nascent tumor cells to B cells through their
immune response to
cells, including T antigens that are
cells, B cells, and not expressed in
Macrophages
NK cells normal tissue
NK cells

NK = natural killer.
1. Norvell A. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:11–24.
 Some Tumor Cells Express Multiple Antigens
That Are Not Expressed by Normal Cells1
Tumor cells release differentially
Normal cells release molecules that are
expressed antigens that cause them to
captured by antigen-presenting cells, but
be recognized as foreign entities and
they don’t elicit an immune response.
therefore elicit an immune response.

NORMAL TUMOR
CELL CELL

1. Finn OJ. N Engl J Med. 2008;358:2704–2715.

 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

TUMOR

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
9
3
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

Activated
T cell

3. Activated T cells migrate back to the

tumor.

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
 T cells Are Important in the Ability of the Immune
System to Detect and Destroy Tumor Cells1

1. Antigens are released by tumor cells and

captured by dendritic cells.
Dendritic cell

Naive T cell

TUMOR 2. Dendritic cells activate

naive T cells in lymph
nodes.

LYMPH
NODE

Activated
4. T cells kill tumor cells through T cell
the release of lytic enzymes or
induction of apoptosis.

3. Activated T cells migrate back to the

tumor.

1. May KF Jr et al. In: Prendergast GC et al. Cancer Immunotherapy. 2nd ed. Elsevier; 2013:101–113.
9
5
 T-Cell Activity Is Regulated By Immune
Checkpoints to Limit Autoimmunity1
Dendritic cell

Inactivated

-
T cell

-
TUMOR
Checkpoints

Checkpoints
Checkpoints
Inactivated LYMPH
T cell NODE

Immune checkpoints, such as

CTLA-4, PD-1, LAG-3, and TIM-3
function at different phases in Activated
T cell
the immune response to
regulate the duration and
level of the T-cell
response.
CTLA-4 = cytotoxic T-lymphocyte antigen 4; PD-1 = programmed cell death protein 1; LAG-3 = lymphocyte activation gene 3;
TIM-3 = T-cell immunoglobulin and mucin protein 3.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–264.
9
6
 Exploiting the PD-1 Immune Checkpoint Pathway

Priming Phase of Effector Phase

Activation

Dendritic cell Naïve T cell Activated T cell Tumor cell

• Emerging research has

PD-1 identified PD-1 as an
immune checkpoint
pathway that tumor cells
may exploit to evade
Antigen
immune surveillance
• Tumor cells may block
immune responses via the
TCR MHC PD-1 immune checkpoint
pathway by expressing the
dual PD-1 ligands, PD-L1
and PD-L2
PD-L1

PD-1
Adapted with permission from Pardoll DM.1 PD-L2

PD-1 = programmed cell death protein 1; TCR = T-cell receptor; MHC = major histocompatibility complex; PD-L1 = programmed
cell death ligand 1; PD-L2 = programmed cell death ligand 2.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–64.
 Exploiting the PD-1 Immune Checkpoint
Pathway
Priming Phase of Effector Phase
Activation
Dendritic cell Naïve T cell Inactivated T cell Tumor cell

-
• PD-1 is upregulated on
PD-1 activated T cells during
the effector phase of the
immune response
Antigen • PD-L1 and PD-L2
engage the PD-1
TCR MHC receptor on T cells to
downregulate T-cell

-
activity in the effector
PD-L1 phase
PD-1

PD-L2

T cell Tumor cells

Adapted with permission from Pardoll DM.1 Inactivation evade destruction
PD-1 = programmed cell death protein 1; PD-L1 = programmed cell death ligand 1; PD-L2 = programmed cell death ligand 2.
1. Pardoll DM. Nat Rev Cancer. 2012;12:252–264.
 PD-1 Pathway and Pembrolizumab
• PD-1 is a negative co-stimulatory
receptor expressed primarily on
activated T cells1
• Binding of PD-1 to its ligands
PD-L1 and PD-L2 inhibits effector
T-cell function1
• PD-L1 expression on tumor cells and
macrophages suppresses immune
surveillance and permits neoplastic
growth2
• Pembrolizumab is a humanized
monoclonal IgG4 antibody
– Binds to PD-1 with high affinity
– Prevents PD-1 from binding to PD-L1
and PD-L2
– Robust antitumor activity and
manageable in multiple advanced
malignanciesa

aFDA-approved for the treatment of patients with unresectable or metastatic melanoma and disease progression following ipilimumab and, if BRAF V600 mutation
positive, a BRAF inhibitor.
1. Keir ME et al. Annu Rev Immunol. 2008;26:677-704; 2. Pardoll DM. Nat Rev Cancer. 2012;12:252-64.
Anti PD-1
 PD-L1 and PD-L2 Can Be Expressed on
Some Tumor Cells
 PD-L1 is expressed on some tumor cells and may
downregulate tumor-specific T-cell activity by binding to
PD-1 in the tumor microenvironment1–5
 – In some tumors, high PD-L1 expression based on immunohistochemistry
 has been associated with a poor prognosis6–8

 PD-L2 may also play a role in helping tumor cells evade

the immune response8–11
 The role of immunology in cancer, including PD-1 and
its dual ligands PD-L1 and PD-L2, is a significant
focus in oncology research
PD-L1 = programmed cell death ligand 1; PD-L2 = programmed cell death ligand 2; PD-1 = programmed cell death protein 1.
1. Hino R et al. Cancer. 2010;116:1757–1766; 2. Wintterle S et al. Cancer Res. 2003;63:7462–7467; 3. Konishi J et al. Clin Cancer Res. 2004;10:5094–
5100; 4. Hamanishi J et al. Proc Natl Acad Sci U S A. 2007;104:3360–3365; 5. Inman BA et al. Cancer. 2007;109:1499–1505; 6. Loos M et al. Ann Thorac
Surg. 2011;91:1025–1031; 7. Thompson RH et al. Cancer Res. 2006;66:3381–3385; 8. Nomi T et al. Clin Cancer Res. 2007;13:2151–2157; 9. Ghebeh H
et al. Neoplasia. 2006;8:190–198; 10. Chapon M et al. J Invest Dermatol. 2011;131:1300–1307; 11. Karim R et al. Clin Cancer Res. 2009;15:6341–6347
Anti T lymphocyte : CTLA4- Ig recombinant protein

MHC class II T cell activation :  Antigen generates ( signal 1)

TCR
 CD28 co-stimulation provides (signal 2)
T cell

APC T Cell Effect

Antigen
Signal 1
CD40 CD40-L +
PD-1L PD-1 -
ICOS-L ICOS +
CD80/86 CTLA4 -
Activated
T cell MHC TCR +
CD80/86 CD28 +
CD80/86CD28
PD-2L ? ?
Signal 2 B7-H3 ? ?

DC = dendritic cell
Moreland (2004).www.medscape.com
 Metastatic NSCLC Patient Flow (United States)1
RG2

ALK, anaplastic lymphoma kinase; CT, computed tomography; EGFR, epidermal growth factor receptor;
MED ONC, medical oncologist; NSCLC, non-small cell lung cancer; PCP, primary care provider; PS, performance status.
1. MK-3475 Spring 2014 Brand Book.
Slide 103

RG2 The information on slides 5-7 was developed from internal market research across the markets of US, EU, and Japan. It is our
understanding that in other regions, such as across AP, practices generally follow those of US or Japan. Please advise if you would like
additional information incorporated to fully capture the markets you need.
Rebecca Goldstein, 12/8/2014
5-Year Survival by Stage of Diagnosis
60%
49%
50% 45%

40%
Percentagea

30% 31%
30%

20%
14%
10% 5%
1%
0%
Stage IA Stage IB Stage IIA Stage IIB Stage IIIA Stage IIIB Stage IV

Disease Stage at Diagnosis

aIncludes NSCLC and small-cell lung cancer. The overall 5-year survival for small-cell lung cancer (6%) is lower than that for NSCLC (18%).2

1. American Cancer Society. NSCLC survival by stage. http://www.cancer.org/cancer/lungcancer-non-smallcell/detailedguide/non-small-cell-lung-cancer.

Accessed September 19, 2014. 2. American Cancer Society. Cancer facts and figures. http://www.cancer.org/acs/groups/content/@research/documents/webcontent/acspc-042151.pdf.
Accessed September 19, 2014.
BIOMARKER TESTING IN NSCLC
 Pembrolizumab is a High-
Affinity, Humanized IgG4, Anti-PD-1
Antibody

• Dual blockade of PD-L1 and PD-L2 --No cytotoxic (ADCC/CDC) activity

• Pharmacokinetics support dosing every 2 weeks (Q2W) or every 3 weeks (Q3W)
• Low occurrence of anti-drug antibodies, which have no impact on pharmacokinetics
• Demonstrated clinical activity in multiple tumor types1-7
• Recently approved in the United States for the treatment of patients with unresectable
or metastatic melanoma and disease progression following ipilimumab and, if BRAF V600
mutation positive, a BRAF inhibitor
1. Ribas A et al. J Clin Oncol. 2014;32(suppl 5):abstr LBA9000; 2. Rizvi N et al. J Clin Oncol. 2014;32(suppl 5): abstr 8007; 3. Garon EB et al. J Clin Oncol. 2014;32(suppl
5):abstr 8020; 4. Seiwert TY et al. J Clin Oncol. 2014;32(suppl 5):abstr 6011. 5. Plimack E et al. Abstr. LBA23. Presented at 2014 ESMO Congress, September 26-
30, Madrid, Spain. 6. Moskowitz CH et al. Bood. 2014;124(21):abstr 290; 7. Nanda R et al. Abstract 1349 (S1-09) presented at SABCS 2014, Dec 9-13, San Antonio, TX.
What is a biomarker? What are the key types of
biomarkers?
• A biomarker is ‘a characteristic that is objectively measured and evaluated
as an indicator of normal biological processes, pathogenic processes or
pharmacological responses to a therapeutic intervention’

Role of the Description of use

biomarker
Predictive or • Used to identify patients who will benefit from a
patient particular drug/therapy
selection
Prognostic • Used to determine how aggressive the disease
process is and/or how a patient may expect to
fare regardless of therapy

Diagnostic • Used to diagnose a disease, potentially before it

is detectable by conventional methods
 Frequency of mutations and availability of
targeted therapies in NSCLC
Gene Alteration Frequency in NSCLC
AKT1 Mutation 1%
ALK Rearrangement 3–7%
BRAF Mutation 1–3%
DDR2 Mutation ~4%
EGFR Mutation 10–35%
FGFR1 Amplification 20%
HER2 Mutation 2–4%
KRAS Mutation 15–25%
MEK1 Mutation 1%
MET Amplification 2–4%
a

NRAS Mutation 1%
PIK3CA Mutation 1–3%
PTEN Mutation 4–8%
RET Rearrangement 1%
ROS1 Rearrangement 1%
a

Lovly C, et al. Molecular Profiling of Lung Cancer. My Cancer Genome. 2016. Available at: https://www.mycancergenome.org/content/disease/lung-cancer/ (accessed June 2016).
PD-L1 AS BIOMARKER IN NSCLC
Detection of PD-L1 by Immunohistochemistry (IHC)

2 Anti-mAb Ab conjugated to
enzyme

1 FFPE tissue section

Anti-PD-L1 mAb 3 Substrate for
enzyme
Deposition of
pigment

Examples of Tumor PD-L1 Expression Scoring:

PD-L1 negative PD-L1 1% PD-L1 70%
 Prospectively defined PD-L1 subgroups

10x
magnification

40x
magnification

Analytically ROC defined

defined cut- cut-off (50%
off (1% TPS) TPS)
Blue stain: Hematoxylin; Brown stain: PD-L1 IHC.
Dolled-Filhart et al. ASCO 2015.
 When to test for PD-L1?
Locally advanced or
metastatic NSCLC
Stage IIIB/IV
DIAGNOSIS
Non-squamous cell Squamous
cell

TREATME EGFR ALK EGFR or ALK

positiv positive negative
NT e
CHOICE
EGFR ALK Platinum based doublet Platinum doublet
First line
inhibitor inhibitor (± bevacizumab) (excluding
pemetrexed)
PD-
L1
Progression Response Response Progression

Maintenance Maintenance
Pemetrexed* gemcitabine or erlotinib
PD-
Progression L1

Second Single agent: Taxane, gemcitabine or EGFR inhibitor (EU)

and third
line
*Only in patients. with non-squamous histology. Clinical trials
 How to test for PD-L1
What happens following tissue extraction? What is the pathology workflow?

Cut Tissue Diagnosis FFPE

sections extraction

Gross Result H&E

FFPE examination

Fixation Dissection IHC Assessment

 Investigational Biomarkers in Lung
Cancer
• Biomarker research in NSCLC still dominated by oncogenic driver mutations, many with low
frequency1
Incidence1-2 Investigational therapiesa
Biomarker
(%) in NSCLC3

KRAS 25 None specifically targeting KRASb

• Crizotinib
• Tivantinib
cMET 4 • Cabozantinib
• PF-02341066
• INC280
RET rearrangement or • Vandetanib
2
translocation • Cabozantinib
• Vemurafenib
BRAF and/or MEK 1
• Dabrafenib and/or trametinib
• Crizotinib
ROS1 1
• Ceritinib
• BKM120
PIK3CA 1
• GDC-0941
• Trastuzumab
HER2/neu 2 • Neratinib
• Afatinib Image from Blakely CM, Bivona TG. Cancer Discov. 2012;2(10):872-875.
aNonexhaustive; many therapies have effects on multiple kinase pathways; some therapies are approved for HER2, human epidermal growth factor receptor 2; MEK, mitogen
other indications but are investigational in NSCLC.1-2 bThere is some evidence that KRAS mutation is predictive extracellular signal-regulated kinase; PIK3CA, phosphatidylinositol-4,
of response to EGFR inhibitors in NSCLC as it is in CRC.2 5-bisphosphate 3-kinase, catalytic subunit alpha.
1. Blakely CM, Bivona TG. Cancer Discov. 2012;2(10):872-875.2. Gerber DE et al. Am Soc Clin Oncol Educ Book.
2014:e353-e365. 3. www.clinicaltrials.gov. Accessed September 3, 2014.
 PD-L1: Investigational Biomarker
for anti–PD-(L)1 Immunotherapy
PD-L1 Expression by NSCLC
• Overexpressed in NSCLC Correlates With Poor Therapy Outcome5
and has potential to be used as a
biomarker1-3
PD-L1 (+) PD-L1 (–)
1.0
• Associated with poor prognosis1-
3
0.8
• Associated with higher response
rates to

Overall Survival
0.6

anti–PD-(L)1 therapy1
0.4
• PD-L1 IHC remains the only
near-term companion diagnostic 0.2

option for drugs targeting in the

PD-1 pathway4 0.0

0 10 20 30 40 50
Month After Surgery
IHC, immunohistochemistry; PD-1, programmed cell death 1; PD-L1, programmed cell death ligand 1.

1. Gandhi L et al. Presented at: AACR 2014; April 6, 2014. Abstract CT105. 2. Zhang Y et al. Onco Targets Ther. 2014:7(12):567–573.
3. Chen Y-b et al. Tumori. 2012;98(6):751-755. 4. Sznol M, Chen L. Clin Cancer Res. 2013;19(5):1021-1034. 5. Mu C-Y et al. Med Oncol. 2011;28(3):682-688.
 TERIMA KASIH

SURABAYA
 Current Immunotherapeutic
strategies in clinic or clinical trials

• Antibody Therapy
• Cytokine Therapy
• Adoptive Therapy
• Vaccination
• Combinational therapy
 Examples of FDA-approved cancer
immunotherapeutic agents
Modality Principle
Hematopoietic stem cell transplantion
(e.g., leukemia and myeloma) 1. Reset the immune system
2. Allo-antigen response
(graft versus tumor
effect)
Antibody (e.g., retuximab, trastuzumab) 1. Eliminate cancer cells
2. Block key signaling
pathways
Cytokines (e.g., type I interferon, interleukin-2) Boost both innate and adaptive
immunity
Dendritic cells (e.g., Sip-T for prostate cancer) Enhance tumor-specific T cell
priming
T cell checkpoint blockade (e.g., Ipilumimab
for melanoma) Block/reverse immune
tolerance
Microbes (e.g., BCG for the transitional
bladder cancer) Enhance innate and adaptive
BREAST Ca
Extrinsic tumor suppression by the immune system

Jeremy B. Swann. J. Clin. Invest. 2007;117:1137–1146

CONCLUSION

•Immunotherapy may be the next great hope for

cancer treatment.
• While antibodies, cytokines, and vaccines have
individually shown some promise, it is likely that
the best strategy to combat cancer will be to
attack on all fronts.
• The effect of immunotherapy in combination with
traditional cancer therapies is another avenue.
Anti PD-1
 Could the Immune System be an
Important Ally in the Fight Against Cancer?
Tumor
• The presence of immune cells
(such as T cells) within a tumor has
been correlated with better clinical T cells
outcome in some tumor types1–3

– Less advanced stage

– Absence of signs of metastasis
– Increased survival
– Reduced risk of relapse
• When the immune system is suppressed, the risk of
developing certain cancers increases
– HIV patients4
– Transplant patients treated with immunosuppressants5
HIV = human immunodeficiency virus.
1. Hwang WT et al. Gynecol Oncol. 2012;124:192–198. 2. Pages F et al. N Engl J Med. 2005;353:2654–2666. 3. Loi S et al. J Clin Oncol. 2013;31:860–
867. 4. Engels EA et al. Int J Cancer. 2008;123:187–194. 5. Grulich AE et al. Lancet. 2007;370:59–67.
Linkage between tumorigenesis, the DNA damage response
and the immune response

Stephan Gasser. Cancer Res 2006; 66(8): 3959-62

Tumor Antigens Eliciting T-Cell Immunity When Presented to Naive T Cells by Antigen-
Presenting Dendritic Cells

Olivera J. Finn, N Engl J Med 2008;358:2704-15

Impact of stress in tumor growth
and the anti-tumor immune
response
Stress alters T-cell profile in tumor
bearing animals
Stress results in increased tumor size
Stress induces mdr1 expression
and drug resistance
 Response to stress
Stress

HPA
Axis
CRF
The nervous, endocrine and immune systems form the
core of an adaptation mechanism to exogenous or
endogenous stresses