(157a) Protein Diffusion, Convection and Vesicular Transport in the Intestinal Wall: How Ramkrishna's Teachings Helped the Drug Delivery Field

The combination of
materials design and advances in nanotechnology has led to the development of
new therapeutic protein delivery systems. The pulmonary, nasal, buccal and
other routes have been investigated as delivery options for protein therapy,
but none result in improved patient compliances and patient quality of life as
the oral route. For the oral administration of these new systems, an
understanding of protein transport is essential because of the dynamic nature
of the gastrointestinal tract and the barriers to transport that exist. Models
have been developed to describe the transport between the gastrointestinal
lumen and the bloodstream, and stimuli-sensitive complexation hydrogels have
been investigated as a class of promising carriers for oral delivery. However,
the need to develop models that accurately predict protein blood concentration
as a function of the material structure and properties still exists. The development of novel carriers and
improved drug delivery profiles requires a multi-pronged approach to fully
assess the viability of improving the absorption and transport of therapeutic
agents. Models allow for the analysis of many iterations before proceeding to
development and experimentation, which can reduce operating costs. Mathematical
models can also be used in conjunction with laboratory experiments to quantify
transport rates and investigate correlations to the animal and human scale.
Cell culture remains one of the most valuable and widely used laboratory
techniques for investigating protein absorption and transport.

The goal of
controlled drug delivery devices is to achieve the desired therapeutic effect
over a long period of time. With traditional tablets or injections, the drug
levels in the blood rise after administration and then decrease until the next
one. A problem with this profile is that sometimes the drug levels in the blood
either achieve toxic levels or ineffective levels. In controlled drug delivery
systems designed for long term administration, the drug level in the blood
follows a profile in which the drug level remains constant, between the
effective maximum and minimum, for an extended period of time. Sustained
released systems are more patient friendly and also offer the advantages of
increased effectiveness and cost effectiveness. The mucosal layers of the body
provide access to the bloodstream and are often marked by high surface areas
for absorption. The use of transmucosal delivery has many advantages, however
each layer presents unique challenges for the delivery of proteins. The
following sections are a review of some of these areas and how they have been
used in the delivery of the previously described therapeutic proteins.

Several approaches
have been used for the study of drug permeation, including transport through
isotropic, lipophilic phases or animal studies. When screening for permeation
characteristics, the choice of test system always represents a compromise
between high throughput with low predictive potential and low throughput with
high predictive potential. Cell cultures correspond to the intermediate level
of complexity of these permeation studies.

Approaches to the
study of drug transport and permeation involve transport through preparations
of intestinal tissue such as everted sac, everted rings, and intestinal loops,
among others. However, these preparations have several disadvantages such as
the lack of cellular metabolism. This may limit their utilization in the study
of active transport processes. Also, membrane isolation processes may damage
the membrane’s potential to carry out enzymatic or carrier functions.

The
two primary pathways for molecules to cross the epithelium and reach the
bloodstream are the paracellular route and transcellular route. Paracellular
transport involves the movement of the molecule of interest between the
epithelial cells whereas transcellular transport occurs when the molecule
passes through the epithelial cell. Hydrophilic and charged molecules primarily
use the paracellular route; however, the size constraints presented by the
tight junctions limit this route to molecules of a size less than 11 A. This
means that only small molecules can pass through the tight junctions. Most
drugs cannot pass freely unless the tight junctions are disrupted.

Three
mechanisms that utilize the transcellular route exist. The first of these
mechanisms is transcellular passive transport. This mechanism is often reserved
for small, neutral molecules that pass through the layer via diffusion without
hindrance. The second mechanism, transcytosis, occurs when a molecule in the
proximity of the cell membrane is entrapped in a lipid bilayer known as a
vesicle. The vesicle passes into the cell and will either release its contents
through the basolateral side of the cell, achieving transcellular transport, or
release it within the cell for digestion. Carrier-mediated transport is the
third transcellular mechanism and occurs when a molecule reversibly binds to
complexes in the lipid bilayer. The complex dissociates within the cell and the
molecule passes to the basolateral side, forms a new complex with the bilayer,
and exits the cell.

The role of
membrane transporters, such as P-glycoproteins, has recently been an important
factor when studying the oral bioavailability of drugs. The elucidated
mechanism of these apically polarized efflux transporters consists of the drug
detection and expulsion after their entrance to the plasma membrane.
P-glycoproteins interact with some antibiotics, cancer chemotherapeutics,
hormones, and surfactants, among others.

The selection of
the cell line is very important in order to mimic the biological barrier with
an in vitro cell culture system. Different human colon carcinoma cell lines
(i.e. Caco-2, HT-29, SW116, LS147T, SW-480) have been considered for the screening
of different drugs. However, Caco-2 and HT-29 have been the most investigated
because of their ability to express morphologic features of mature enterocytes
or goblet cells.