The changing face of therapeutics

Question 1

What are monoclonal antibodies?

Answer 1

members of the Ig superfamily
IgG 1-4 with different constant domains

Question 2

Which mab is mainly used in therapies

Answer 2

IgG1 or 3
IgG 1 has good antibody binding

Question 3

How many types of monoclonal antibodies are there?

Answer 3

4 types
Murine (-omab)
Chimeric (-Ximab)
Humanised (-zumab)
Human (-umab)

Question 4

What are the mechanisms of therapeutic antibodies?
Give an example drug for each mechanism

Answer 4

Cytokine & growth factor blockade
Ligand blockade
Receptor blockade
Receptor downregulation
Depelting and signalling antibodies
Weaponised antibodies
Bispecific antibodies

Question 5

Neutralising and blocking antibodies

Answer 5

Ligand blockade: infliximab - a recombinant humanized monoclonal antibody hand binds soluble TNF alpha
Receptor blockade: Tocilizumab - a recombinant humanized monoclonal antibody IL-6 receptor inhibitor used to treat autoimmune conditions
Receptor downregulation: Efalizumab - a recombinant humanized monoclonal antibody binds to CD11 receptors on T lymphocytes

Question 6

Depleting and signalling antibodies:

Answer 6

Antibody-dependent cell mediated cytotoxicity (ADCC)
Antibody-dependent cellular phagocytosis (ADCP)
Complement-dependent cytotoxicity (CDC)

Question 7

Explain the neonatal fragment crystallisable (Fc) receptor FcRn

Answer 7

Antibodies (e.g. IgG and albumin) have long plasma half lives
Ingested via pinocytosis
Once ingested they bind to the neonatal fragment crystallisable receptor (FcRn)
But only binds to the FcRn when its acidic (pH 5.0-6.5) [not in its physiological pH 7.4]
acidification = FcRn binds IgG (and albumin)
Retained within endosome
FcRn bound IgG os transported back to the plasma membrane
Vesicle exocytosis =IgG is released into circulation

Question 8

What is Myasthenia Gravis?

Answer 8

Autoimmune disease caused by IgG against alpha 1 subunit of the nicotinic acetylcholine receptor in neuromuscular junctions

Causes loss of motor activity
Initial stages treat with cholinesterase inhibitors
No disease modification

Question 9

State a drug treatment for Myasthenia Gravis

Answer 9

Efgartigimod (VYVGART)

Question 10

Efgartigimod MOA

Answer 10

FC Frgament (pH independent to FcRn) > binds to receptor > Lowers IgG and blocks IgG transfer across barriers

Question 11

Weaponised antibodies (oncology)

Answer 11

-Radioactive element delivers a short range burst of radiation to cells targeted by the antibodies

or

-Chemical linkage to a bacterial toxin

Question 12

Bispecific antibodies

Answer 12

Multiple functionally different binding domains that allow for interaction with two target antigens

Question 13

State the strengths and weaknesses of antibodies

Answer 13

Strengths:
High success rate
Well tolerated
Generally specific (less off target effects)

Limitations:
Expensive with high production costs
Limited penetration to CNS
Cannot be administered orally

Question 14

What are the benefits of using nanobodies?

Answer 14

Robust binding molecules (more potent)
Chemical and thermal stability
Better penetration into tissues and organs

Question 15

State the types of dysfunctional proteins

Answer 15

Poor translation
Truncated protein
Protein that doesn’t go where its supposed to
Protein does not function
Excess protein

Question 16

What is spinal muscular atrophy (SMA)

Answer 16

SMN1 encodes for survival motor neurone protein 1 - essential for nerve maintenance
Sufferers inherit two copies of a missing or faulty (mutated) SMN1 gene

Motor neurones wither and die

Question 17

DNA therapeutics (Vectors)

Answer 17

Replacing therapeutic proteins when delivered to the nuclei of patients cells

Vectors > lentivirus, adenovirus
Eludes the hosts immune defences
(Viruses are self-inactivating and replication-deficient)

Question 18

Genomic editing

Answer 18

Adding, removing or altering genetic material

Utilize nucleases are guided by DNA binding proteins or RNA to cleave genomic DNA

Zinc finger nucleases
Transcription activator-like effector nucleases (TALENs)
Clustered regularly interspaced short palindromic repeats (CRISPR) and associated proteins (Cas)

Question 19

What are the 2 different Gene editing systems?

Answer 19

Clustered regularly interspaced short palindromic repeats-CRISPR = single-guide RNA carries DNA cutting enzyme Cas9 to target sites

Transcription activator-like effector nucleases -TALEN = TALEs are fused with the DNA scissors, Fok1

Question 20

RNA therapeutics +example
(mRNA delivery)

Answer 20

SARS-COV2 Vaccine
Pfizer/Biontech = mRNA for spike protein in oily capsule (nanoparticle)

Question 21

What are Antisense Oligonucleotides?
What are the 2 types?

Answer 21

short single stranded DNA phosphorothioate DNA, RNA analogs, complementary to a certain region of RNA they are meant to target

-Alter RNA and reduce, restore or modify proteins expression

2 types:
RNase H-dependent ASO-provides (good for excessive protein = knockdown)

RNase H-independent ASO-provies steric block = changes expression of proteins by inhibiting/preventing splicing machinery

Question 22

Exon skipping

Answer 22

Alisense oligonucleotides can target splice sites leading to exon skipping or shifting the ratio between existing splice forms

Question 23

Small interfering RNAs (siRNAs)

Answer 23

Small non-coding RNA duplexes that originate from precursor siRNAs
Effective at silencing gene products

Question 24

Premature stop codon

Answer 24

• Nonsense mutations result in loss of function characteristics

Drugs can prevent premature stop codons
E.g. in cystic fibrosis readthrough agents can be used

Question 25

Stem cells (Extra learning ??)

Answer 25

Remove induced pluripotent stem cells (IPSCs)
treat with gene editing technology
Apply DNA alkylating agent for the dividing cells
Infuse with modified stem cells

Sickle cell aneamia treatment

Question 26

What are antisense oligonucleotides?

Question 27

What is CRISPR/CAS9