Nanomedicines + EPR

Question 1

What happens to drugs with a log p <1.7?

Answer 1

They are incorporated into the aqueous compartment (Hydrophilic).

Question 2

What happens to drugs with a log p >5?

Answer 2

They are retained in the lipid bilayer (Hydrophobic)

Question 3

What happens to the drugs with an intermediate log p- between 1.7 and 5 and why is it an issue?

Answer 3

This is the majority of drugs. They undergo compartment partitioning- a component into the aqueous compartment and another component into the lipid compartment
- This is an issue because it makes it easier for the drug to leave the liposome entirely

Question 4

How are liposomal drugs normally made?

Answer 4

• The lipid components (and the drug if it is lipid soluble) are added to the organic solvent and freeze-dried. This forms a ‘Lipid cake’.
• Aqueous solution is added including water-soluble drug.
• This is then hydrated with further water and mixed (Agitation)
• This forms large, multi-lamellar (multiple lipid bilayers) vesicles
• To sperate into individual vesicles (single layer), requires sonification, extrusion and homogenisation techniques.

Question 5

What are the 3 classifications of liposome vehicles?

Answer 5

Small unilamellar vesicles (SUV): 25-100 nm
Large multilamellar vesicles (LUV): 100nm- 1 micrometre)
Multi-lamellar vesicles (MLV): >1 micrometre

• Is based on the size and number of lamellae

Question 6

What is the advantage of adding cholesterol to liposomes?

Answer 6

• Cholesterol can be made up to 50% of the total lipid- this means the liposome has an extended planar group. This is very hydrophobic and so occupies the internal tail group of the lipid bilayer = Making the bilayer stiffen and rigid
  = This rigidity leads to a decrease in permeability of the bilayer and so increases drug retention inside the liposome

Question 7

What is most commonly added to liposomes as surface modifications?

Answer 7

PEG chains- Polyethylene glycol chains as part of the head groups

Question 8

What is the advantage of adding PEG chains as surface modifications to liposomes?

Answer 8

The addition of PEG chains to the surface acts as a disguise.
The PEGs become the outside of the liposome and aren’t recognised by the body.
- Additionally, the peg chains prevent aggregation of liposomes by acting as steric repulsions

Question 9

What modifications can be made to liposomes and why?

Answer 9

• Adding Cholesterol- can be made up to 50% of the total lipid- this means the liposome has an extended planar group. This is very hydrophobic and so occupies the internal tail group of the lipid bilayer = Making the bilayer stiffen and rigid
  = This rigidity leads to a decrease in permeability of the bilayer and so increases drug retention inside the liposome
• The addition of PEG chains to the surface acts as a disguise.
  The PEGs become the outside of the liposome and aren’t recognised by the body.
• Additionally, the peg chains prevent aggregation of liposomes by acting as steric repulsions
• Can attach antibodies to the outside of the liposomes- allows specific recognition of a target site

Question 10

How can you prevent the drug from partitioning out of the liposome prior to the target site?

Answer 10

By making the liposomes via Remote drug loading:
- Make the liposome without the drug
- In the aqueous component of the liposome, some kind of gradient is needed such as pH, ionic strength or the addition of a gelating agent or something that will bind to the drug within the liposome
- When the drug is then added to the liposome, the drug is captured by one of the methods mentioned above until the liposome is delivered to the target site.

Question 11

What kind of compound can be added to the aqueous compartment of remote drug loading to keep the drug in the liposome until target site is reached?

Answer 11

E.g. Ammonium Sulphate
- This binds to the drug to form a complex within the liposome so that the drug has to stay within the liposome

Question 12

What are the reasons for encapsulating a drug in a liposome?

Answer 12

• It can alter the pharmacokinetic properties and bio-distribution of the drug. 50% of the time it is the ADMET (Absorption, distribution, metabolism, excretion and toxicology) properties that cause a drug to fail at clinical trials, but when a drug is enclosed in a liposome, it is the properties of the liposome that matter and not the drug itself!
• Can function as a drug reservoir- slowly releases the drug
• Protection of the drug from enzyme attack in the body e.g. peptide and nucleotide-based drugs need to be protected from DNAase and Peptidases

Question 13

What are the 2 drug examples that you need to know for this topic?

Answer 13

• Doxorubicin (Myocet)
• Doxorubicin ( Caelix/Doxil)

Question 14

What are some problems with drugs used for tumours and solutions by using liposomal drug delivery systems?

Answer 14

• Poor solubility - can cause the drug to precipitate. SOLUTION- These delivery systems can provide both hydrophobic and hydrophilic environments and so drug solubility can be enhanced
• Tissue damage due to extravasation-Regulation of the drug release can reduce or eliminate tissue damage
• Rapid breakdown of drug in vivo leading to loss of activity- can protect the drug or form sustained release preparations
• hAS UNFAVOURABLE PHARMACOKINETIC PROPERTIES, MEANINGg RAPID CLEARANCE and needing higher doses- can change the pharmacokinetics of the drug, reduce the clearance and renal clearance of small drugs is avoided
• Lack of selectivity for the target tissues- leads to low uptake by the therapeutic target and causing side effects in other tissues- EPR can aid in ligand targeting = more specific

Question 15

How do lipsomal drugs end up in the MPS depot?

Answer 15

• When IV drugs are administered, blood opsonins mark them for phagocytosis (destruction) as they are foreign cells
• The opsonins recognise cell-like structures, so the larger the liposome, the more cell-like it is and its lipid composition and surface charge all mimic a cell = recognition
• The liposomes that get opsonised, enter the mononuclear phagocytic system (MPS)
• This leads to the accumulation of the drug-containing liposomes at these sites. The MPS acts as a drug depot and the drug is slowly released into circulation

Question 16

What does MPS stand for?

Answer 16

Mono-nuclear phagocytic system

Question 17

What increases a liposome’s chance of being opsonised?

Answer 17

• Larger size
• Lipid composition
• Surface charge

Question 18

If you don’t want a liposome to enter the MPS system, what do you need to do to it?

Answer 18

• Need to AVOID opsonisation = modify the stability, clearance rate and tissue distribution of the liposomes
• This is done by changing the surface characteristics e.g. adding PEG chains to the outside of the liposomes ( Stealth liposomes- the peg coating is inert and so avoid opsonins and can reach the target site). This creates hydrophilic surfaces that repel opsonins and maintain liposomes in circulation.

Question 19

What does EPR stand for?

Answer 19

Enhanced permeation and retention

Question 20

What is the liposome formulation of the Myocet form of doxorubicin?

Answer 20

• It is enclosed in an egg-phosphatidyl choline/cholesterol liposome
• It is a LUV liposome - 180 nm diameter = large vehicle
• As it is a large vehicle, it is more cell-like and it has no PEGylated layer (no disguise) = so will be recognised by opsonins
• Is therefore accumulated within the MPS system and slowly released into the body

Question 21

What is Myocet used for?

Answer 21

1st line breast cancer treatment (Is doxorubicin)

Question 22

What is the liposome formulation of the Caelyx form of Doxorubicin?

Answer 22

• The doxorubicin is formulated into liposomes containing a DSPE PEG 1000 coating
• They are SUV liposomes (<100 nm diameter)
• Therefore, as they are small and have a PEG disguise, they are able to avoid opsonisation and can persist in circulation in blood = Accumulate at the tumour site via EPR effect

Question 23

What is Caelyx used for?

Answer 23

Breast cancer
Ovarian cancer
Kaposi’s sarcoma

Question 24

What is the formulation of albumin-bound paclitaxel and how does it work?

Answer 24

Albumin-bound paclitaxel is used in the treatment of breast cancer and non-small-cell lung cancer
- It uses human serum albumin as the nanoparticle- as albumin is a normal component of our blood (30-50 g/L) the drug system is recognised as self- is self-non-antigenic
- The albumin encapsulates the hydrophobic drug (paclitaxel) and produces a hydrophilic particle that is taken up by transcytosis via GP60 glycoproteins= it therefore exploits a natural cellular response and can easily cross the cell membrane

Question 25

Where have nanoparticle technologies been recently majorly useful?

Answer 25

In the development and production of Covid-19 vaccines- e.g. Astrazeneca and pfizer vaccines

Question 26

What is the definition of clearance?

Answer 26

The volume of blood plasma which is completely cleared of a substance (drug) per unit time.

Question 27

What are the 2 main mechanisms of clearance of a drug?

Answer 27

Renal - in urine
Hepatic- Bile –> faeces

Question 28

What determines whether a drug is renally or hepatically cleared?

Answer 28

The SIZE of the drug:
- Small particles can be filtered by the glomerulus in the kidney ( below 6-8nm diameter)
- Larger particles remain in the blood and then the mononuclear phagocytic system (MPS)- here they are retained for months to years and are slowly released
- If too large for renal clearance and avoids MPS e.g. due to PEG coating, it will be hepatically cleared

Question 29

What is the problem with drugs ending up in the MPS for years/months?

Answer 29

• It can be a safety issue as they accumulate and can increase the presence of toxic side effects.

Question 30

Why is there caution when using large liposomes for doxorubicin drugs?

Answer 30

Because large drugs can be retained in the MPS system for along time and doxorubicin has a long-term dose-dependent risk of cardiotoxicity.

Question 31

What is the difference between passive and active targeting in nanomedicine?

Answer 31

• Passive = Deposition of nano-sized systems within the tumour microenvironment is enhanced due to characteristics inherent to the tumour- not manipulating the body in any way
• Active- manipulating the exterior of the nanoparticle by adding antibody-targeting proteins- these bind to the therapeutic target in the body. Is not in the clinic yet!

Question 32

What usually happens to small molecules in the body and why does using nanomedicines help?

Answer 32

• Small molecules are poorly targeted and are rapidly excreted in the urine = therefore few reach their target and some accumulate in irrelevant tissues = unwanted side effects
• Nanomedicines are cleared much slower, so they are in the body longer and can travel to their therapeutic site and accumulate. Less are released into irrelevant tissues = decreased side effects

Question 33

Why do traditional cancer therapies cause so many side effects?

Answer 33

Because they are not specific- so can’t Dtarget certain cells and instead act on all tissues causing unwanted side effects.

Question 33

Discuss the permeability aspect of the EPR effect in chemotherapies?

Answer 33

• In a tumour, the angiogenesis (blood vessels) are erratic- they are irregular and can become leaky and breakdown
• This allows the material to easily leave the blood and enter the tumour tissue = enhanced permeability at the desired site

Question 33

Discuss the retention aspect of the EPR effect in chemotherapies?

Answer 33

• In a tumour, the lymphatic system is broken down. This means that the lymph system isn’t draining away material like usual and so it accumulates in the TME. Thos means drugs stay at the target site for longer and so have an enhanced therapeutic effect.