1. IMMUNE CHECPOINTS & IMMUNOTHERAPY IN CANCERS
HUỲNH QUYẾT THẮNG
VIETNAM CANCER SOCIETY

2. INHIBITORY IMMUNE CHECKPOINTS IN CANCERS
Negative regulators of immune system
Maintaining self-tolerence
Preventing auto-immunity
Blocking immune checkpoint is a promising approach for activating anti-tumor immunity . It reactivate the auto-immunity

3. The immune checkpoint proteins in immune cycle

4. CTLA-4

5. CTLA-4: Cytotoxic T lymphocyte associated protein 4
FUNCTIONAL STRUCTURE:
Known as CD 152, an homolog of CD 28
Functioning as an IC, down regulating immune responses
Both bind with CD80 & CD86 (also called B7-1 & B7-2): ON & OFF switches on APC
FUNTIONAL STRUCTURE OF IMMUNE CHECKPOINT CTLA-4

6. CTLA4: Functional structure
FUNCTIONS
Member of Ig. superfamily, expressed by activated T.cell and transmits a negative signal to T cell (reverse to CD28)
CTLA4 binds to CD80 &86 with greater affinity & avidity than CD28 → out-compete for its ligands
Also found in regulator T cell, contributing to inh. Function.
Activating of T cell leads to increase expression of CTLA4
CTLA4: Functional structure

7. CTLA4 &homolog.CD28 bind to B7 1 và 2
FUNCTIONS (cont.):
Mechanistically, CTLA4 recruits a Phosphatase for TCR →attenuating signal
Recent suggests in vivo: CTLA4 capture & remove B7-1&2 from APC → unavailable trigger CD28.
DC -Treg interaction→ sequestration of Fasin-1, essential for synapse formation and skew fascin -1-dependent actin polarization in AP(D)C → adhesion zone → reduced T cell priming.
Also function via modulation of cell motility &/or signaling through PI3 kinase.
CTLA4 &homolog.CD28 bind to B7 1 và 2

8. CTLA4:
T.Cell motility appears to give evidence for the so-called: “Reverse stop signaling model”from TCR.
CTLA4 reverses “TCR induced stop signal”, for firm contact between Tc & APC

9. CTLA4
STRUCTURALLY:
Extracellular domains , trans-membrane & cytoplasmic tail.
Alternative splice variant, encoding different isoforms, have been characterized by a disulfide bond, while the soluble isoform functions as monomer
The intracellular domain ia similar to CD28: No intrincic catalytic activity, contains YVKM motif binding PI3K; PP2A & SHP2 & one proline rich motif binding SH3 containing protein
KEY ROLES:
Inhibit T cell responses
Affect signaling, via competing with CD28
Also binds PI3K

10. CLINICAL SIGNIFICANCE
Variants in this gene associated with:
Diabetis I, Grave’s disease, Hashimoto, SLE
Polymorphism of CTLA4 are associated with:
Autoimmune diseases
Germline Haploinsufficiency
CHAI’s disease
Clinical and laboratory manifestation
Abnormalities of immune system:
- Agonist to reduce immune activity: Potential therapy for autoimmune disease.
- Antagonist to increase immune activities: possible : Potential therapeutic against cancers

11. CLINICAL SIGNIFICANCE OF CTLA4

12. BLOCKADE OF CTLA4 AND CANCERS
Antibody againts CTLA4 (Ipilimumab) treat melanoma 2011
Tremelimumab (not yet approved
Antigen specific activating T CD4+ is the key initiation of adaptive immunity.
The native T cell require 2 signals:
- Antigen specific interact TCR with MHC
- Co-stimulator, arisen through the surface of CD28, stimulate the interaction with B7 on APC

13. PD-1/PDL1-2

14. Programmed cell death Protein 1, also known as CD279
PD-1:
Programmed cell death Protein 1, also known as CD279
Cell surface receptor, down-regulates the immune system, promoting “self tolerence”
An IC and guards against autoimmunity via a dual mechanism: promoting apoptosis in antigen specific T cell; Reducing apoptosis in regulatory T cells.
A new class of drug blocking PD-1: activates the immune system to attack the tumor → to treat some cancers

15. PD-1 Structure Membrane protein of 286 AA.
Member of extended CD28/CTLA4 family of T cell regulators
Includes an extra cellular IgV domain, transmembrane & intracellular tail containing two phosphorylation sites located on inhibitory motifs, suggesting that PD-1 negatively regulates TCR signals
PD-1 is expressed on surface of T cell, B cell & macrophage

16. PD-1 has two ligands, PD-L1 & PD-L2, members of family B7
Protein PD-L1 upregulates on marcrophage & DC in response to LPC, GM-CFS treatment; on T cell, BC & TC Receptors signaling.
PD-L1 is expressed on almost all murine tumors cell lines: PA1, P815, B16
PD-L2 expression is more restricted, expressed mainly on DC and a few tumor lines.
PD-1: PD-Ligands C

17. FUNCTION OF PD-1
Protein PD-1 in humans encoded by PDCD1, a cell surface receptor of Ig superfamilty, expressed on T cell & Pre-B, binds two ligands PD-L1 và PD-L2
PD-1 & ligand negatively regulate immune responses: PD-1 develops glomerulonephritis on knockout mice; cardio-myopathy on C57BL/6 & BALB/c bacgrounds. In vitro, treatment of anti-CD3, stimulated T.cell by PD-L1 Ig, result in decrease of T cell proliferation và IGF-γ secretion.
It’s suggested that Tcell CD8+ is more susceptible to inhibition by PD-L1.
PD-L1 – PD-1 interaction inhibits activation, expansion & acquisition of effector function of virus-specific CD8+, which can be reversed by blocking PD-1/PD-L1 interaction.

18. MECHANISMS OF ACTIVITIES
FOR CTLA4 &PD-1
CTLA negatively regulates the antitumor immunity, PD-1 is explored as a target of immunotherapy.
The expressed PD-L1 on tumor cell inhibits the antitumor activity of PD-1 on effector T cell.
Initiation of PD-1 via PD-L1 leads to production IL-10 → T CD4

19. CLINICAL SIGNIFICANCE
CANCER
PD-L1 highly expresses in cancers, PD-1 has the crucial role in immune evasion.
Inhibitory interaction between PD-1 and PD-L1 blocks the immunity control .
Combining TLA4 and PD-1 antibodies is an striking inhibitory immune checkpoint
The combination of PD-1 & CTLA4 A.bodies has been shown to be more effective than either
A. body alone.
PD-1 A.B reactivates the ability of CD8 Tcell to lyse cancer cell
PD-L1 expression is not absolute determinant of therapy effectiveness
FDA has approved a combination Nivolumab & Ipilimumab 10/2015

20. ANTI - PD-1
THERAPY
A number of cancer immunotherapy targeting PD-1 receptor has been developed: Nivolumab (opdivo-BMS) produced complete/partial responses in NSCLC, melanoma, renal-cell cancer, colon and pancreatic cancer did’nt have response
Nivolumab targets PD-1 receptor was approved in Japan on 7/2014 & on 12/2014 in USA-FDA to treat metastatic melanoma.
Pembrolizumab (Keytruda. MK 3475 Merck) also targeting PD-1 receptor, was approved by FDA on 10/2015 for NSCLC progressed after other treatments .
Other drugs targeting PD-1: Pilidizumab (Cure Tech),BMS
Atezolizumab (MPDL 3280A Roche), Avelumab (Merck…) target PD-L1.

21. FUTURE PROSPECTS
The clinical successes have opened up a new area of therapeutic: the FDA approval of anti-CTLA4 , quickly followed by anti-PD-1 therapy.
A new found awareness of potential antitumor activity of patient’s endogenous immune systemự once the BRAKE have been released have clarified the role of ICs (determined by genetic & biologicalanalyses ) in immune responses, specially in different levels.
The exploration of iCs bring two chalenges:
Definition of potential bio-markers that can determine which IC pathway dominate in particular tumor is inhibitor of choice? Is this possible that specific oncogenic pathway as PI3K-AKT hay STAT3 may induce and can be used as surrogate biomarker?
The clinical development of combinational approaches: Vaccin associated with anti-IC ?

22. CONCLUSIONS
Clinical results CTLA4 & PD-1/PD-L1 inhibitors suggests that B7-H1/PD-1 pathway is an important target for the effective antitumor immune response.
Favor and toxicity profile of PD-1 & B7-H1 inhibitors verify that the large usefullness for the treatment of advanced cancers.
The possible prolongation of response for the inhibition of PD-1 & CTLA4 pathways until 10 years, leading to extend globally the clinical trials on the spectrum of solid tumors
Many other complex approaches include blokade of immune checkpoints associated with new vaccine, Angiogenesis, MAPK inhibiting targeted therapy.