Interferons

Question 1

3 distinct classes of interferon

Answer 1

• IFN-α
• IFN-β
• IFN-γ

Question 2

Interferon

Answer 2

A substance secreted by virally infected cells that mediate resistance

Question 3

Functions of interferons

Answer 3

• Induction of cellular resistance to viral attack
• Regulation of immune function
• Regulation of growth and differentiation of many cell types
• Sustenancce of early phases of pregnancy in some animal species

Question 4

Type I interferons

Answer 4

• Acid-stable

IFN-α & IFN-β

• 30% amino aicd identity
• bind to same receptor
• induce similar activities

Question 5

Type II interferon

Answer 5

IFN-γ

• evolutionarily distinct from other interferons
• binds to separate receptor
• induces different range of activity

Question 6

Biological effects of type I interferons

Answer 6

Antiviral activity
Anti-proliferative effect

Question 7

Anti-tumour effects of type I interferons

Answer 7

Have ability to increase NK and T-cytotoxic cell acitivity

Question 8

Biological effects of type II interferons

Answer 8

Weak antiviral and anti-proliferative activity.
Represents main macrophage-activating factor

IFN-γ

Question 9

Macrophage-mediated effects

Answer 9

• Destruction of invading microbes
• Destruction of intracellular pathogens
• Tumour toxicity
• Increased MHC antigen expression leading to enhanced activation of lymphocytes via antigen presentation

IFN-γ

Question 10

What happens when IFN-γ is co-administered with type I interferons

Answer 10

Potentiates the antiviral and anti-proliferation activities of IFN-α & IFN-β

Question 11

Initial therapeutic application of interferons

Answer 11

• Identified in late 1950s but therapeutic application rendered impractical due to low levels produced by the body.
• Large scale purification from sources like blood was non viable.
• Interferons exhibit species preference - only human derived interferons can be used in human medicine

Question 12

First production of interferon in small quantities

Answer 12

• Until 1970s
• Sourced from human leukocytes obtained from transfused blood supplies
• only 1% pure
• contained mixture of various IFN-αs

Question 13

First production of interferon in large quantities

Answer 13

• late 1970s
• mammalian cell culture
• cancer cell lines were found to secrete greater quantities of interferons and were amenable to large scale culture due to their transformed nature

Question 14

Namalaw cell line

Answer 14

A specific strain of human lymphoblastoid cells
- the major industrial source of interferon

Question 15

Interferon production from namalwa cell line

Answer 15

• Cells propagated in animal cell fermenters
• The additon of an inducing virus (sendai virus)
• Results in production of leukocyte interferon.
• Final product contained 9 IFN-α subtypes

Question 16

Tradename given to one of first approved interferon products

Answer 16

Wellferon

Question 17

Most interferons currently in medical use

Answer 17

Recombinant human products produced by E. coli

Question 18

Clinical Studies of IFN-α

Answer 18

• Conducted in the late 1970s
• Found that IFN-α could induce regression of tumors in breast cancer, certain lymphomas, and multiple myeloma.
• Could also delay the recurrence of tumor growth after surgery in patients with osteogenic sarcoma.

Question 19

First recombinant interferon to become available for clinical studies

Answer 19

IFN-α2a (Intron A)
In 1980

Question 20

Schering Plough’s rhIFN-α2b (Intron A)

Answer 20

First approved in 1986 in US for hairy celll leukaemia - now 50+ countries

Now approved in the US for hepatitis B and C

Question 21

What is the producer microbe for Intron A and where is it expressed?

Answer 21

• E. coli.
• Expressed intracellularly and harbors a cytoplasmic expression vector (KMAC-43) containing the interferon gene.

Question 22

Where are the manufacturing facilities for Intron A located and how is the processing undertaken?

Answer 22

• New Jersey and in Brinny, Co. Cork.
  1. The upstream processing (fermentation) and downstream processing (purification and formulation) are physically separated, being undertaken in separate buildings.
  2. Fermentation is undertaken in 42,000L stainless steel vessels.
  3. After recovery, a number of chromatographic purification steps are undertaken within a large cold-room adapted to function under cleanroom conditions.
  4. Crystallization of the IFN-α2b is then undertaken as a final purification step.

Question 23

Modified recombinant IFN-αs

Answer 23

PEGlyated interferons

• PEG Intron A
• ViraferonPeg

Sythetic interferons

• Infergen

Question 24

ViraferonPeg

Answer 24

• PEGylated
• Approved EU since 2000
• Differs by presence of covalently attached PEG
• Schering Plough
• Treats chronic hepatitis C
• Displays prolonged half-life compared to non-PEG
  (13-25 hours compared to 4 hours)

Question 25

How does the biological activity of PEGylated interferons compare to non-PEGylated interferons?

Answer 25

Essentially the same. Half life different: * PEGylated 13-25hrs * non-PEGylated 4hrs

Question 26

What is the advantage of using PEGylated interferons?

Answer 26

The PEGylated product displays a significantly prolonged plasma half-life (13-25 hrs, compared to 4 hrs for the unpegylated species).

Question 27

Infergen

Answer 27

* Interferonalfacon-1 or consensus interferon * Engineered interferon * treats hepatitis C * Development entailed inital sequence comparisions between a range of IFN-αs * Amino acid sequence reflects most frequently occurring amino acid residue in each corresponding position of these native interferons

Question 28

Medicinal uses of IFN-β

Answer 28

treatment of relapsing-remitting multiple sclerosis (MS)

Question 29

IFN-β prepartions approved for use

Answer 29

* Betaferon * Betaseron * Avonex * Rebif

Question 30

How does IFN-β mediate its affect

Answer 30

Inhibiting the production of IFN-α & IFN-γ; thus mediating down-regulation of the pro-inflammatory response

Question 31

What is the typical reduction in relapse frequency observed with IFN-β-based drugs in multiple sclerosis (MS) patients?

Answer 31

IFN-β-based drugs reduce the frequency of relapses by approximately 30%.

Question 32

Medicinal use of IFN-γ

Answer 32

Treatment of chronic granulonatous disease (CGD) a rare genetic disorder

Question 33

CGD Chronic granulonatous disease

Answer 33

* CGD is a genetic defect in the mulitcomponent NADH oxidase system. * This system produces oxidative species that are lethal to pathogens

Question 34

Traditional treatment of CGD

Answer 34

Prophylactic administraion of antimicrobial agents in attempt to prevent occurrence of infection

Question 35

Why is prophylactic admin. not favourable?

Answer 35

* Destroys the microbiota creating a resevoir for antimicrobial resistance for immunocompromised patients * Creates major public health risk * Creates environment for superbugs to evolve

Question 36

Recombinant human IFN-γ used therapeutically

Answer 36

* Produced in E.coli * Termed IFN-γ1b * Displays same bio activity but lacks carbohydrate compound

Question 37

IFN-γ1b product

Answer 37

Tradename: Actimmune Manufactured: Genetech

Question 38

Actimmune clinical trial results

Answer 38

* Reduction in the incidence of life threatening infections by > 50% * Reduction in the incidence of total infections by > 50% * Reduction in the number of days of hospitilisaiton by 3-fold

Question 39

How does IFN-γ work?

Answer 39

Functions as a potent activator of phagocytes, improving their ability to generate toxic oxidative products (via the NADPH oxidase system)

Question 40

Additional function of IFN-γ

Answer 40

Also promotes increased expression of IgGFc receptors on the surface of phagocytes – thus increasing the phagocyte’s ability to destroy opsonized infectious agents.

Question 41

What are some conditions that IFN-γ shows promise for treating?

Answer 41

Treatment of: * Leishmaniasis caused by the parasitic protozan of the genus Leishmania, * Lepromatous leprosy caused by Mycobacterium leprae.

Question 42

Additional interferons

Answer 42

* IFN-τ (trophoblastin) * IFN-ω (omega)

Question 43

IFN-ω (omega)

Answer 43

* 170 aa glycoprotein * Exhibits 50-60% aa identity to INF-αs * Even more closely related to IFN-τ. * Found in humans, pigs and other mammals. * A recombinant form of feline IFN-ω has been approved for the reduction of parvoviral infections in young dogs.

Question 44

IFN-τ (trophoblastin)

Answer 44

* Type I - but exhibits lower toxicity * Amino acid sequence and functional similarities with IFN-α * can promote inhibition of reverse transcriptase activity in cells infected with HIV * 3 - 4 functional genes in sheep & cattle