First Aid, Chapter 4 Laboratory Tests, Hybridoma and Monoclonal Antibodies

Question 1

What is a hybridoma?

Answer 1

A hybridoma is product of cell fusion between a normal antibody producing B lymphocyte and a myeloma cell, followed by selection of fused cells that secrete antibody of the desired specificity.

Question 2

How are hybridomas made?

Answer 2

• Spleen cells from an immunized mouse are fused with myeloma cells by using polyethylene glycol (PEG) to produce a hybrid cell line called a hybridoma.
• Myeloma cells lacking both antibody production and the enzyme hypoxanthine-guanine phosphoribosyl transferase (HGPRT) are selected out. Absence of this enzyme prevents growth in hypoxanthine-aminopterinthymidine (HAT) medium.
• After fusion, cells are transferred to HAT medium, where unfused spleen and myeloma cells die. Only hybrid cells with HGPRT enzyme (i.e., gene contributed by spleen cells) can grow in this medium and are, thus, selected.
• Hybridomas are screened for antibody production, and cells that make antibody of desired specificity are cloned by growing them from a single cell (Figure 4-22).
• As each hybridoma is descended from a single cell, all the antibodies it produces are identical in structure, including their antigen-binding site and isotype. These antibodies are called monoclonal antibodies (mAbs).

Question 3

What is the mneumonic for monoclonal antibodies?

Answer 3

Mnemonic XImab: miXing human and mouse makes Chimeric
Umab: hUman
Zumab: humans at the Zoo are humaniZed
Omab: mOuse

Question 4

What type of antibodies are -omabs? How are they synthesized? What is the murine component?

Answer 4

Type: mouse

Means of synthesis: Murine-variable and constant antibody regions

Murine component (%): 100%

Question 5

What type of antibodies are -ximabs? How are they synthesized? What is the murine component?

Answer 5

Type: chimera

Means of synthesis: chimera

Murine component (%): 30-35%

Question 6

What type of antibodies are -zumabs? How are they synthesized? What is the murine component?

Answer 6

Type: humanized

Means of synthesis: Combines murine mAb hypervariable or CD regions to remaining portion of human IgG molecule

Murine component (%):

Question 7

What type of antibodies are -umabs? How are they synthesized? What is the murine component?

Answer 7

Type: human

Means of synthesis: Human heavy- and light-chain variable (V) region genes are cloned into a bacteriophage—V region is expressed as a fusion protein on phage cell surface → phage-display library—multiplied in bacteria-select phage with desired antigen specificity. May replace hybridoma technology for production of mAbs

Murine component %: none

Question 8

How are -cept proteins synthesized?

Answer 8

Fusion proteins created by joining two or more genes that were originally coded for separate proteins

Question 9

Describe the nomenclature of product source identifiers in monoclonal antibodies.

Answer 9

u=human
o=mouse
a=rat
e=hamster
i=primate
xi=chimeric
axo=rat/mouse
xizu=combination of humanized and chimeric chains

Question 10

What is the target antigen for abciximab? What is the application?

Answer 10

Target antigen molecule: Integrin αIIβ3

Application: Cardiac ischemic complications

Question 11

What is the target antigen for adalimumab? What is the application?

Answer 11

Target antigen molecule: TNFα

Application: Rheumatoid arthritis

Question 12

What is the target antigen for alemtuzumab? What is the application?

Answer 12

Target antigen molecule: CD52

Application: Chronic lymphocytic leukemia

Question 13

What is the target antigen for basiliximab? What is the application?

Answer 13

Target antigen molecule: IL-2 receptor α chain (CD25)

Application: Transplant rejection

Question 14

What is the target antigen for belimumab? What is the application?

Answer 14

Target antigen molecule: B-cell activating factor

Application: Systemic lupus erythematous (SLE)

Question 15

What is the target antigen for bevacizumab? What is the application?

Answer 15

Target antigen molecule:
Vascular endothelial growth factor (VEGF)

Application: Colorectal cancer

Question 16

What is the target antigen for brentuximab vedotin? What is the application?

Answer 16

Target antigen molecule: CD30

Application: Anaplastic large-cell lymphoma (ALCL), Hodgkin’s lymphoma

Question 17

What is the target antigen for canakinumab? What is the application?

Answer 17

Target antigen molecule: IL-1B

Application: Cyropyrin-associated periodic syndrome (CAPS)

Question 18

What is the target antigen for cetuximab? What is the application?

Answer 18

Target antigen molecule: Epidermal growth factor receptor

Application: Colorectal cancer, head and neck cancer

Question 19

What is the target antigen for certolizumab pegol? What is the application?

Answer 19

Target antigen molecule: TNFα

Application: Crohn’s disease

Question 20

What is the target antigen for daclizumab? What is the application?

Answer 20

Target antigen molecule: IL-2 receptor α chain (CD25)

Applicaton: Transplant rejection

Question 21

What is the target antigen for denosumab? What is the application?

Answer 21

Target antigen molecule: RANK ligand

Application: Postmenopausal osteoporosis, solid tumor’s bony metasteses

Question 22

What is the target antigen for Eculizumab? What is the application?

Answer 22

Target antigen molecule: Complement system protein C5

Application: Paroxysmal nocturnal hemoglobinuria

Question 23

What is the target antigen for Efalizumab? What is the application?

Answer 23

Target antigen molecule: CD11a

Application: Psoriasis

Question 24

What is the target antigen for Gemtuzumab? What is the application?

Answer 24

Target antigen molecule: CD33

Application: Acute myelogenous leukemia

Question 25

What is the target antigen for Golimumab? What is the application?

Answer 25

Target antigen molecule: TNF-α

Application: Rheumatoid arthritis, psoriatic arthritis, Ankylosing spondylitis

Question 26

What is the target antigen for Ibritumomab tiuxetan? What is the application?

Answer 26

Target antigen molecule: CD20

Application: Non-Hodgkin’s lymphoma

Question 27

What is the target antigen for Infliximab? What is the application?

Answer 27

Target antigen molecule: TNF-α

Application: Several autoimmune disorders

Question 28

What is the target antigen for Ipilimumab (MCX-101)? What is the application?

Answer 28

Target antigen molecule: CTLA-4

Application: Melanoma

Question 29

What is the target antigen for Keliximab? What is the application?

Answer 29

Target antigen molecule: CD4

Application: Multiple sclerosis

Question 30

What is the target antigen for Mepolizumab? What is the application?

Answer 30

Target antigen molecule: IL-5

Application: Hypereosinophilic syndrome

Question 31

What is the target antigen for Muromonab-CD3 (OKT3)? What is the application?

Answer 31

Target antigen molecule: T cell CD3 receptor

Application: Transplant rejection

Question 32

What is the target antigen for Natalizumab? What is the application?

Answer 32

Target antigen molecule: α 4 chain of integrin molecule

Application: Multiple sclerosis

Question 33

What is the target antigen for Ofatumumab? What is the application?

Answer 33

Target antigen molecule: CD20

Application: Chronic lymphocytic leukemia

Question 34

What is the target antigen for Omalizumab? What is the application?

Answer 34

Target antigen molecule: FcεRI (cε3)

Application: Asthma

Question 35

What is the target antigen for Palivizumab? What is the application?

Answer 35

Target antigen molecule: RSV F-protein

Application: Infants with bronchopulmonary dysplasia

Question 36

What is the target antigen for Panitumumab? What is the application?

Answer 36

Target antigen molecule: Epidermal growth factor receptor

Application: Colorectal cancer

Question 37

What is the target antigen for Ranibizumab? What is the application?

Answer 37

Target antigen molecule: Vascular endothelial growth factor A (VEGF-A)

Application: Macular degeneration

Question 38

What is the target antigen for Ritiximab? What is the application?

Answer 38

Target antigen molecule: CD20

Application: Autoimmune diseases, B-lymphocyte lymphomas

Question 39

What is the target antigen for Tocilizumab? What is the application?

Answer 39

Target antigen molecule: Anti-IL6R

Application: Rheumatoid arthritis

Question 40

What is the target antigen for Tositumomab? What is the application?

Answer 40

Target antigen molecule: CD20

Application: Rheumatoid arthritis

Question 41

What is the target antigen for Trastuzumab? What is the application?

Answer 41

Target antigen molecule: ErbB2

Application: Breast cancer

Question 42

What has reduced the risk of anaphylaxis with monoclonal antibodies?

Answer 42

There is a risk of hypersensitivity reactions, ranging from mild allergic reaction to anaphylaxis. However, with the development of humanized and human mAbs, the risk of hypersensitivity has decreased significantly by reducing exposure to murine antibody immunogenicity.

Question 43

Describe how fusion proteins are made?

Answer 43

Fusion proteins are created by joining two or more genes that originally coded for separate proteins.

• Translation of the fusion gene results in a single polypeptide with functional properties derived from each original protein.
• Recombinant fusion proteins are created artificially using recombinant DNA technology for use in research and therapeutics.

Question 44

Give an example of a fusion protein. What is it made form? What is it used for?

Answer 44

One example is etanercept, a TNFα antagonist created through the combination of a tumor necrosis factor receptor (TNFR) with the immunoglobulin G1 Fc segment. TNFR provides specificity for the drug target, and the antibody Fc segment adds stability and deliverability of the drug. It is used for treatment of autoimmune conditions, including rheumatoid arthritis, psoriatic arthritis, ankylosing spondylitis, plaque psoriasis, and juvenile idiopathic arthritis.