Lecture 9 Flashcards
(97 cards)
1
Q
What causes B cell activation?
A
T helper cell
Activation is through the MHC-II/ peptide interacts with the T helper cell with the specific TCR
2
Q
What causes T helper cell activation?
A
same pathogen is picked up by the other antigen presenting cells – macrophages and dendritic cells
–> presented to the naïve T cells
–> T helper cell with TCR that recognizes
3
Q
How does class switching of B cells occur?
A
- Activated Th cells interact with B cells via CD40 ligand
–> B cells proliferate, differentiate and secrete antibodies
2.The cells (Th) secrete cytokines that determine class switching
4
Q
How do Th activate B cells?
A
- interact with B cells via CD40 ligand
—> B cell proliferate, differentiate and secret antibodies - release cytokins that determine class switching
5
Q
How do CD8 T cells get activated?
A
- cytokines (IL-2 and IL-21) from CD4 T cells
- stimulation from APC through IL-12 and IL-15
6
Q
What cytokines from CD4 T cells activate CD8 T cells?
A
IL-2 and IL-21
7
Q
Are the antigen peptides presented on the MHC-I and MHC-II molecules same or different?
A
Different
8
Q
What were the ‘magic bullets’ from Paul Ehrlich?
A
- mAb linked to a toxin (e.g. ricin)
- mAb linked to radioactive compound
- mAb to induce ADCC (antibody dependent cellular cytotoxicity)
9
Q
Why are mAbs not very ‘useful’?
A
- can’t activate human effector functions
- Formation of immune complexes –> elimination
- induction of allergy
10
Q
What is mouse antibody called?
A
omab
11
Q
What is human antibody called?
A
umab
12
Q
What is chimeric antobody and what is it called?
A
Ximab
The 4 variable regions are from mouse and all the constant regions are from human
13
Q
What is abciximab and what is it for?
A
variable regions from mouse +constant regions from human
Against CD41/CD61 antigen
14
Q
Abciximab
What is CD41/CD61 antigen for?
A
–> expressed exclusively on platelet
–> receptor for fibrinogen, lead to platelet activation and aggregation, lead to blood clot
15
Q
What is Glanzmann thrombasthenia and what is it caused by?
A
- mutation on gene for CD41/CD61
-bleeding disorder due to defective clot formation
16
Q
When is abciximab usually administered?
A
non-surgical procedure of percutaneous coronary angioplasty
17
Q
What is the other name for CD41/CD61 antigen?
A
GPIIb/IIIa
18
Q
Apart from CD41/CD61 antigen, what else can convert fibrinogen (insoluble) to fibrin (soluble)?
A
Thrombin (made from prothrombin through the action of clotting factors)
19
Q
What are polyhedrocytes?
A
Angular erythrocytes held together by platelets
20
Q
What is the first mAb approved for clinical use?
A
Abciximab
21
Q
What happens after the administration of abciximab?
A
platelets level drops but returns to normal level after 120hr
22
Q
What is infliximab for?
A
binds to TNFα
23
Q
Infliximab
What is TNFα?
A
–> pro-inflammatory cytokine that produce an acute phase reaction.
–> can kill cancer cells
–>expressed as membrane bound trimer, released as soluble form
24
Q
Infliximab
What is TNFα produced by?
A
activated macrophages
CD4+ lymphocytes
NK cells
neutrophils
mast cells
eosinophils
neurons
25
How infliximab (anti-TNFα) administered?
intravenous injection
26
What is the function of TNFα druring inflammation?
released by macrophages
--> induce vasoldilation
--> expression of E-selection and P-selection to slow down leukacytes
-->promotes expression of proteins that trigger blood clotting
27
What happens if release of TNFα becomes systemic?
massive vasodilation --> drop in bp (septic shock) --> organ failure and death
28
What is Crohn's disease and what is it treated by?
Inflammation in the digestive tract
--> bleeding
--> holes in intestinal wall (perforation)
--> fistulas
Treated with infliximab, which is anti-TNFα
29
What is Ulcerative colitis?
an inflammatory bowel disease (IBD) that causes long-lasting inflammation and ulcers (sores) in the innermost lining of your large intestine (colon) and rectum
30
What is a fistula?
A fistula is an abnormal connection between an organ, vessel, or intestine and another organ, vessel or intestine, or the skin
31
What can fistula between urethra and rectum lead to?
urinary tract infection
32
What is rheumatoid arthritis and what is it treated with?
Autoimmune disease
--> inflammation of swelling of joints
Treated with infliximab
33
What is rheumatism?
any disease marked by inflammation and pain in the joints, muscles, or fibrous tissue, including rheumatoid arthritis.
34
According to Neils Jerne, what are the three antibody libraries?
1. immune library
2. universal library
3. engineered library from natural and synthetic Ab gene sequences
35
What is immune library?
From blood samples of individuals who had contracted a particular disease or had been vaccinated against it.
-->proved to have worked
--> contain affinty matured antibody
-->can also be obtained from primates colely related to humans
36
What is universal library?
from natural naïve human antibody genes.
--> broad and unbiased specificities
--> obtainable from B cells of non-immunized donors
37
What is hybrid natural/ synthetic libraries?
They can be constructed on an antibody framework, with engineered one or more CDRs in the heavy and/or light chains.
38
How are engineered antibodies produced?
The cDNA of the heavy and light chains can be inserted into plasmid vectors that can
support expression.
39
What is the selection of eukaryotic cells for expression of plasmid DNA for antibodies based on?
1. support the folding of the combined heavy and light chains
2. A secretion system
40
Name 3 cells lines that are used to produced engineered antibodies through plasmid expression
1. Chinese Hamster Ovary (CHO)
2. Human Embryonic Kidney (HEK-293)
3. COS-7 (derived from Monkey kidney fibroblasts)
41
In engineered antibodies, what is the function of a 'linker'?
A linker may be used to replace the constant
regions of the heavy and light chains in scFv
--> made up of AAs
-->ensure the 2 trimeric CDRs are together
42
What is scFv?
single chain variable fragment
-->single-chain with variable domains from the heavy and light chains
43
What AA does the 'linker' consist of? Why?
GGGGS-GGGGS-GGGGS
G4S linker ( consist of repeats of GGGGS)
G (Glycine) for the flexibility of linker
S (Serine) to provide solubility
44
What is the purpose of glycine (G) in linker>
flexibility of linker
45
What is the purpose of serine in linker?
to provide solubility
46
What is the size of the linker for scFv?
~15 AAs
47
What structure can scFv exist as?
Diabody (~5AA) --> 2 antigen binding sites
Triabody (<5AA) --> 3 ABS
Tetrabody (<5AA) --> 4ABS
48
What is the 'specialty' of engineered Trastuzumab?
scFv
--> inserted monovalent red fluorescent protein 1 (mRFP1), used for detection of Her2 receptor for breast cancer that are Her2 positive
--> also used as drug
49
When linking 2 Ig variable domains, what should be paid attention to?
The CDRs need to face the same side
50
How does size affect the therapeutic effect of mAbs?
- Small size for short retention time due to renal elimination
-Good penetration to tissues such as solid tumors
51
How does valency affect the therapeutic effect of mAbs?
Monovalent: short retention time
Bivalent and higher: enhanced retention time
52
How does affinity affect the therapeutic effect of mAbs?
High affinity = high retention time
High affinity not good for penetration
53
What are bi-specific antibodies?
a class of antibodies engineered to simultaneously bind to two different antigens or epitopes.
54
What is a tandem single chain diabody what are its advantages?
a type of bispecific antibody (VH1VL1 + VH2VL2)
--> improved retention time + possible to engineer bispecific versions
55
What is a minibody?
bispecific mAb with only CH3 domain
56
What is a chemical cross-linked F(ab’)2?
a type of bispecific antibody, does not have Fc for retention by Fc receptors
Fab1+Fab2 (joined together)
57
What are some types of bispecific antibodies?
1. Tandem single-chain diabody
2. Minibody
3. Chemical cross-linked F(ab’)2
4. Bispecific antibody by fusion of two hybridomas
58
What is the BiTE (Bispecific T cell Engager) mAb called?
blinatumomab
59
What is BiTE?
Bispecific T cell Engager
Two scFv with different antigen-binding specificity to bring about a T-cell to the target.
60
What is the specificity of BiTE blinatumomab?
two scFv
one against CD3 on cytotoxic T cells
one against CD19 on tutor cells
61
What is the function of BiTE?
Brings (cytotoxic) T cells to deliver the lethal hit to the tumour cells.
*Note that it also target normal B cells, but their elimination is clinically manageable.
62
Does BiTE have same affinity for both of its target antigens i.e. CD3 and CD19? What purpose does it serve?
assymetric affinity: anti-CD3 (T cell) high and anti- CD19 (cancer) low
--> make the T cell killing in an “enzymatic” manner
63
What cancers could be treated with BiTE (blinatumomab)?
colorectal cancer
gastric adenocarcinoma.
prostate cancer.
64
What is one way of producing bispecific antibodies?
fusion of 2 hybridoma - quadroma
65
In an antibody, what holds the heavy chains together?
constant domains 2 and 3 of the heavy chains (when heavy chain has 3 constant domains)
66
In an antibody, what holds the heavy chain and the light chain together?
constant domain 1 of the heavy chain and the constant domain of the light chain
67
When forming quadromas through fusion of hybridomas in a bid to produce bispecific antibodies, what challenge has to be overcome?
Want:
light chain A-heavy chain A-heavy chain B-light chain B combination
However, 8 different combinations can be formed, need to eliminate their formation
68
Which isotope of Ig usually preferred for forming quadromas through fusion of hybridomas?
IgG1
69
In CH3 of IgG1, the AAs at which position are necessary for spatial compatibility?
Y86 <---> T22
T22 <---> Y86
Y=tyrosine
T=threonine
70
During fusion of 2 hybridomas (mAb1 and mAb2), how to ensure that the heavy chain of mAb1 will 'interact' with heavy chain of mAb2 in order to from a bispecific antibody?
through engineering in CH3--> forcing formation
original:
Y86 <---> T22
T22 <---> Y86
Engineering:
mAb1:
Y86 ---- Y22
Y22 ---- Y86
mAb2:
T86 ---- T22
T22 ---- T86
Spatially incompatible to 'interact' with its own kind --> form quadroma
71
For the formation of hybridomas, engineering at CH3 solves the problem of pairing of correct heavy chains (one from each mAb), how is it ensured that the light chain stays with it 'parent' heavy chain? for instance, light chain of mAb-A 'stays' with heavy chain of mAb-A?
exchange the constant region of the light/heavy chains in ONE OF THE mAb
--> EITHER mAb-A OR mAb-2
Possible combination would require the
interaction between the CL region of the
light chain, and the CH1region of the heavy chain
72
Name a bispecific IgG1 mAb that utilises the following:
1. “knob-into-hole” mutation in the CH3 domain.
2. Exchange of the CH1 and CL domains of the anti-ANG-2
Faricimab for wAMD
against VEGF-1 and Angiopoietin-2
Also, mutation in the Fc region such that the FcRn binding capacity is removed – so that RG7761 does not stay around for too long.
73
Name some bispecific antibodies used as drugs
1. blinatumomab (BiTE)
2. Faricimab (wAMD)
3. Catumaxomab
4. Lutikizumab (osteoarthritis)
74
Why is the specificity of catumaxomab and what is its mechanism?
1. EpCAm on tumor cell
2. CD3 on T lymphocyte
3. Fc receptors on macrophages, dendritic cell and NKC
The two “antigen-binding” arms bring the tumour cell and T ell together, to promote T cell-mediated killing by lysis (release of perforin) or by induction of apoptosis.
75
What kind of antibody is Catumaxomab?
Oman (mixed mouse and rat)
76
For an fusion of hybridomas from different species, what is the advantage?
ensures the correct pairing the heavy and light chains, as the heavy and light chains from the same species have the preference to combine
77
What is the mechanism of Catumaxomab?
The two “antigen-binding” arms bring the tumour cell and T ell together, to promote T cell-mediated killing by lysis (release of perforin) or by induction of apoptosis.
The Fc part was also engineered to bind to various Fc receptors to recruit the additional arsenal of the immune system in the forms of phagocytes, NK cells, and dendritic cells.
78
What type of mAb is Lutikizumab what is it for?
DVD-Ig
Double variable doman immunoglobulin
Lutikizumab is to treat osteoarthritis
79
What kind of antibodies do cartilaginous fish and camelids have?
single heavy chain antibodies
80
What are cartilaginous fish?
Backbone made up of cartilage
Cartilage (軟骨) is a strong, flexible connective tissue that protects your joints and bones. We have cartilage too.
81
What has has the CD40L and what has the corresponding CD40?
CD40L --> Th cells
CD40 --> antigen presenting cells (B cells and dendritic cells)
82
What are chimeric antibodies?
Chimeric antibodies are a type of engineered monoclonal antibodies that contain regions derived from different species, typically human and non-human sources, to combine the desired characteristics of each.
83
What is ankylosing spondylitis?
Long term inflammation of the spine leading to fusing of vertebrae
--> 85% patients are HLA-B27 positive
--> cured with infliximab
84
What is psoriasis?
Psoriasis is a skin condition that causes flaky patches of skin which form scales.
Treated with infliximab
85
Name some medical conditions that are treated with Infliximab?
1. Psoriasis
2. Rheumatoid arthritis
3. Crohn's disease
4. ankylosing spondylitis
5. Psoriatic arthritis
86
What is otelizixumab?
against the ε chains of the CD3 complex to prevent T cells from killing β pancreatic cells in patients of type-1 diabetes.
87
What mutation do patients of breast cancer have in their genome?
1 in 5 have extra copies of HER2 gene (Human epidermal growth factor)
HER2 can dimerize to signal cell to proliferate
Can form heterodimers with HER1 and HER3
88
What mAb used to treat breats cancer?
Trastuzumab and Pertuzumab
89
What is the mechanism of trastuzumab and pertuzumab in treating cancer?
trastuzumab and pertuzumab: prevent formation of homodimer and heterodimers + recruit NK cells thorugh Fc receptors
--> antibody dependent cell mediated cytotoxicity
90
What is eculizumab and ravulizumab for?
against the complement protein C5
deregulation of complement activation leading to anemia and unwanted blood clotting:
91
What is PNH and what is it cause by?
paroxysmal nocturnal hemoglobinuria
--> deficiency in CD59 which prevents the formation of lytic complex
--> leads to
92
What is aHUS?
mutations in the regulatory proteins in factor H, factor I,
MCP, DAF, and CR
--> lysis of erythrocytes
--> deregulation of C3 leads to blood clots and kidney failure
93
What is Denosumab?
- human IgG2 antibody
-treat osteoporosis
94
What is osteoporosis hmoeostasis acheived by?
RANKL and OPG
The reduced level of estrogen in post menopause (絕經後) women, the production of OPG is suppressed. The balance shifts to the effect of RANKL, which stimulates the osteoclasts to bone resorption, resulting in osteoporosis
95
What is the relationship between RANKL?
RANKL is a cytokine that plays a central role in bone resorption by promoting the differentiation and activation of osteoclasts, which are cells responsible for breaking down bone tissue.
96
What is OPG?
OPG is a soluble decoy receptor for RANKL. It acts as a negative regulator of bone resorption by binding to RANKL and preventing it from interacting with RANK on osteoclast precursor cells.
97
