pegylated liposomes

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PEGylated Liposomes

enhance Bioavailability

(Molecular stealth technology prolongs release and minimizes toxicity and side effects By Will

Block.)

hen we go swimming in the ocean, we are, in a sense, intruders in an alien environment,

one in which we are ill-equipped to survive should anything go wrong. Somewhere in the back of every swimmer’s mind must be the unsettling thought that he or she has entered

the world of … Jaws. Without any protection. For decades, scientists have been trying, without

notable success, to develop effective shark repellents. But forget about repellents—wouldn’t it becool if we could apply some kind of chemical camouflage to our skin so that sharks would see us

merely as big watery blobs rather than something meaty and edible?

Conventional Liposomes Are Good . . .

Right—dream on. As we will see below, however, that dream has been realized with regard to

liposomes, which are microscopic, manmade cells used as sustained-action delivery vehicles for a wide variety of drugs, vaccines, enzymes, nonenzyme proteins, and genetic material—and

now, for some nutritional supplements as well. The molecular “payload” is encapsulated inside

the liposomes, which eventually break down through natural processes and spill their contentsinto the bloodstream or into tissues to which they have migrated by diffusion through the walls

of capillaries. Liposomes are a safe and effective way to introduce agents into our system that are

problematic for some reason when taken orally, as well as agents that cannot be taken orally at

all (because they’re degraded by digestive juices), such as nucleic acids and most proteins.

The problem with conventional liposomes, though, is that they’re seen as alien invaders by

phagocytes, our own built-in version of Jaws. Phagocytes are cell-eating cells whose mission is

to devour anything that doesn’t belong in our bloodstream. And when liposomes are devoured,so are their therapeutic payloads, which may thus go to waste—but not necessarily. If the

payload is not broken down by enzymes inside the phagocyte, it may remain there and perhaps

do some good—provided that it was designed for that purpose. Some drugs are, in fact, designedto treat phagocyte-related diseases or to enhance phagocytic activity. Thus, allowing them to be

gobbled up by phagocytes is perfect.

Another possible outcome in case the payload survives the onslaught of the phagocyte’sdegradative enzymes is that it may escape back into the bloodstream. This phenomenon is calledmacrophage-mediated drug release (a macrophage is one type of phagocyte).1 It provides a

gradual, prolonged release, which is usually a good thing, as we will see below. Since it occurs

primarily in the liver and spleen, where phagocytes are especially abundant, conventional

liposomes can be used to deliver certain drugs for treating diseases of these two organs.

. . . But PEGylated Liposomes Are Better



Generally speaking, though, phagocytes are bad news for liposomes, which they usually dispatch

within minutes of administration.2 Camouflaging the liposomes so as to fool phagocytes into

ignoring them thus became a key objective of pharmaceutical chemists during the 1970s and beyond. (Liposomes were discovered in England in 1965.3) The result of their efforts was a

process called PEGylation, in which countless molecules of a synthetic, nontoxic polymer,

polyethylene glycol (PEG), are attached, at one end of the polymer chain, to the surface of theliposome.4,5 The long, slender, highly flexible PEG molecules slosh around the liposome like

spaghetti boiling in a pot. Because of their chemical affinity for water molecules, they are

heavily hydrated. To phagocytes, this molecular “cloak” of water of hydration makes thePEGylated liposomes look like little watery blobs rather than something edible, so they tend to

leave them alone. Mission accomplished!

The two main advantages of PEGylated liposomes for delivering drugs or supplements areincreased bioavailability and the possibility, in some cases, of targeted delivery to the organs or tissues that most need them.1 Let’s look at these two benefits in turn.

Types Of LIPOSOMES:

There are four basic kinds of liposomes, each with its own advantages and disadvantages. All of

them consist of a spherical cell membrane (lipid bilayer) that encapsulates a payload of

therapeutic molecules. All of them bypass the digestive tract, thus increasing the bioavailability

of their payloads, which remain biologically inert until the cell membrane ruptures. When andwhere that occurs, and why and how it occurs, is what makes the difference. It depends on many

different physical and chemical properties of the liposomes and on the physiological environment

in which they find themselves.

Conventional liposomes are “naked,” i.e., they’re ordinary liposomes made from simple

phospholipids, with nothing to protect them from phagocytes. Their surfaces (both exterior andinterior) are either electrically neutral or negatively charged (depending on the specific nature of

the lipid molecules used in making the bilayer. These liposomes are used mainly to deliver drugsto the phagocytes that eat them, so as to treat disorders of the phagocytes themselves or to treat

diseases of the liver or spleen, where most phagocytes are found. The latter form of treatment is

made possible by drugs that are re-released unharmed by the phagocytes.

PEGylated liposomes are “stealth” liposomes that evade detection and destruction by phagocytes by virtue of their cloaks of hydrated PEG (polyethylene glycol) molecules. Their purpose is two-

fold: (1) to increase the bioavailability of drugs or supplements by bypassing the digestive tract,

and (2) to minimize any potential toxicity or side effects of these agents by remaining in the

circulation for a long time and releasing their payloads slowly. As a bonus, they’re passivelytargeted to tumors and to inflamed tissues, where they’re preferentially absorbed because of the

increased permeability of the capillaries that nourish these tissues.

Immunoliposomes are stealth liposomes that have been specially designed for active targeting toa given type of tissue or organ that the liposome is able to recognize by its molecular fingerprint.

This is accomplished by highly specific chemical modifications to the PEG molecules or the lipid



bilayer of the liposomes, or both. The molecular entity responsible for cell recognition is usuallyan antibody or an antibody fragment. Antibodies are immunoglobulins, which are glycoproteins

produced by the body’s immune system—hence the “immuno” part of the name. These liposomes

are used primarily for cancer therapy.

Cationic liposomes are stealth liposomes whose surfaces (both exterior and interior) are positively charged so as to increase the loading efficiency of their payload, which is recombinant

DNA. These liposomes are used for gene therapy, in which certain disorders are treated by

introducing specifically engineered genetic material (DNA) into the patient’s cells through activetargeting.

PEGylated Liposomes Offer Increased Bioavailability

Throughout the drug and supplement industries, poor bioavailability of therapeutic agents is a

huge problem, especially where older people are concerned.6

For example, as we age, our abilityto absorb certain nutrients from the gut, such as vitamin B12 and folic acid, declines markedly.

Other nutrients, such as the antioxidants resveratrol and quercetin, have poor bioavailability to begin with, because they’re completely metabolized in the gut and liver before they ever reach

the general circulation. The metabolites of resveratrol and quercetin do enter the circulation,

however, and are probably responsible for some of the beneficial effects (observed mainly inlaboratory and animal studies) attributed to the parent compounds. Yet other nutrients, such as

coenzyme Q10, the body’s vital “spark plug” molecule, are so costly that we need to maximize

their bioavailability so we can take lesser amounts and still obtain the benefits.

By keeping the blood concentrationof the drug or supplement low and

relatively constant, we can avoid the

pharmacological yo-yo effect (peaks

and troughs) that has been the norm

with traditional delivery methods.

We can address such problems with PEGylated liposomes, which can be used to deliver nutrients

to the bloodstream via a more direct and less hostile route than the digestive tract, namely, the

mucous membranes of the mouth (or, in some cases, the vagina or labia).* The liposomes, in the

form of aqueous suspensions, can also be applied transdermally (through the skin), but thetransmucosal route seems to be more effective. Once absorbed by the mucosa, the liposomes, if

they’re small enough, enter the bloodstream by diffusing through the exceedingly thin walls of capillaries, whence they travel via the venous system to the heart, and thence via the arterial

system to all the body’s capillaries, to begin the cycle anew (one complete circuit takes about a

minute).



*Liposomes themselves are destroyed by digestive juices, so it’s not possible to sneak liposome-encapsulated nutrients intothe circulation via the digestive tract.

A Long, Slow Release Is Best

In the circulation, the PEGylated liposomes eventually begin breakingdown and spilling their payload, which, until the moment of release, has

remained biologically inert because of its entrapment. For a variety of

reasons, the process is long and slow—it can take many hours or even afew days, versus the relatively short “burst release” that occurs in most

conventional forms of drug or supplement delivery.

By keeping the blood concentration of the drug or supplement low and

relatively constant, we can avoid the pharmacological yo-yo effect (peaksand troughs) that has been the norm with traditional delivery methods,

especially with agents that are rapidly removed from the bloodstream

(within hours or even minutes) by natural processes. Think of a river: for the plants and animals that depend on it, it’s better that the river run

placidly all the time than that it alternate between flooding and drying up.

Bonus: Toxicity and Side Effects Are Minimized

Another advantage of keeping the blood levels of a drug or supplement

low is that it tends to minimize any toxicity or side effects, which areoften concentration-dependent. A case in point is the anticancer drug

doxorubicin, which can cause irreversible heart damage as well as nausea

and vomiting; another is the antibiotic/antifungal drug amphotericin B,

which can cause kidney damage along with lesser side effects. PEGylatedliposomes are now being used to deliver these and many other drugs with

better results than were possible previously.7 (The phospholipids used for making liposomes are harmless, by the way, as is polyethylene glycol;

when the liposomes rupture, the phospholipids may be incorporated into our own cell walls, and

the PEG is excreted.)

PEGylated Liposomes Can Offer Some Targeted Delivery

The other main advantage of PEGylated liposomes lies in the possibility of targeted delivery of drugs or supplements to the tissues or organs that need them most. Not only does this maximize

the delivery efficiency for the agent in question, but it also minimizes the chances of toxicity toother organs, as can occur with doxorubicin, amphotericin B, and many other drugs.

There are two kinds of targeted delivery: passive targeting, which is easy to accomplish butlimited in scope, and active targeting, which is difficult to accomplish but fraught with potential

for many applications.8 As we will see, a key role in targeting of either kind is played by theendothelial cells that line the walls of our blood vessels, including the all-important capillaries.



Passive Targeting Occurs for Tumors and Inflamed Tissues

Passive targeting does not depend on any particular property of the liposome (whether

PEGylated or not) that would help it find its target. Instead, it depends on a particular physical property of two kinds of tissues, namely, developing tumors and inflamed tissues. The capillaries

that nourish these tissues typically show increased permeability (leakiness) compared with thecapillaries found in normal tissues. This phenomenon is called the enhanced permeation and

retention (EPR) effect.6

The leakiness occurs when the tiny gaps between the endothelial cells of the capillary walls are

larger than normal. This makes it easier for liposomes to diffuse through the capillary walls, and

they are therefore preferentially absorbed by these tissues (which is what passive targeting is).

There they eventually break down and deliver their payload. For obvious reasons, the liposomalagents most commonly used for passive targeting are anticancer and anti-inflammatory drugs.

The efficiency of the process depends critically on having liposomes in the correct size range.

Active Targeting Is a Tall Order

Whereas passive targeting arises accidentally from a physical property of certain tissues, activetargeting relies on deliberate chemical modifications of the PEG molecules or the phospholipid

molecules that constitute the cell walls of the liposomes themselves—or both.* The possibilities

for such modifications are nearly infinite, and achieving an optimal combination of manydifferent physical and chemical properties of the PEGylated liposomes is vital for success. It

requires great expertise in multiple scientific disciplines.

*The term PEGylation, coined in reference to the basic PEG molecule with which this stealth technology was begun, hassince come to encompass not just the basic PEG molecule, but also any of the innumerable chemical derivatives of PEG

that have been synthesized by organic chemists to serve specific biological functions of liposomal delivery systems.Polymers other than PEG are also used, and, for that matter, lipids other than phospholipids are also used to makeliposomes. To achieve active targeting, the preferred method is to attach cell-specific antibodies (immunoglobulins) to thefree ends of the PEG molecules.

The objective is to design the physicochemical properties so precisely that the PEGylatedliposomes will have a strong chemical affinity for the cells of a certain kind of tissue or organ in

the body—heart cells, liver cells, eye cells, etc.—as well as the ability to deliver their payloads to

those cells in the right way under the right conditions. Although the PEGylated liposomes cannotseek out target cells, à la the “magic bullet” concept introduced by the great German

bacteriologist Paul Ehrlich a century ago, they can recognize these cells by their unique

molecular “fingerprints” when they encounter them. The idea is that the liposomes will thenattach themselves to these cells (and to no others) and disgorge their contents directly into thecells. It’s a tall order, but it is being accomplished, with increasing success, by pharmaceutical

chemists.

A Highly Sophisticated Infant Technology



The concept of active targeting is stunning, and the technologies that the pharmaceutical

companies have been developing to implement it (of which PEGylated liposomes are just one of

many) are highly sophisticated. They are also massively patent-protected, so the use of activetargeting for nutritional supplements, if any, will probably be tightly controlled by these

companies.7 Although the research on active targeting with PEGylated liposomes has been

underway since the 1970s, the practical applications—all of them with prescription drugs thus far —are still so new that most current textbooks of pharmacology make no mention of them. The

field is still in its infancy, and many exciting developments are surely yet to come, some of

which may spill over into the nutritional supplements arena.

Next month we’ll look at the amazing technology of gene therapy using liposomal delivery of DNA, and the problem of how to get liposomes into the most inaccessible part of the human

body—the brain.

References

1. Molecular stealth technology prolongs release and minimizes toxicity and side effects ByWill Block.

2. Hillery AM, Lloyd AW, Swarbrick J, eds. Drug Delivery and Targeting for Pharmacistsand Pharmaceutical Scientists. Taylor & Francis, London, 2001.

3. Papahadjopoulos D, Allen TM, Gabizon A, Mayhew E, Matthay K, Huang SK, Lee KD,

Woodle MC, Lasic DD, Redemann C, Martin FJ. Sterically stabilized liposomes:

improvements in pharmacokinetics and antitumor therapeutic efficacy. Proc Natl Acad

Sci USA 1991;88:11460-4.

pegylated liposomes

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