A new study by research chemists at the University
of Warwick has challenged a century old rule of pharmacology that defined
how quickly key chemicals can pass across cell walls. The new observations of
the Warwick researchers suggest that the real transport rates could be up to
a hundred times slower than predicted by the century old "Overton's Rule".
This could have major implications for the development and testing of many future
Overton's rule says that the easier it is for a chemical to dissolve in a lipid
(fat) the easier and faster it will be transported into a cell. The Rule was
first outlined in the 1890s by Ernst Overton of the University of Zürich.
He declared that substances that dissolve in lipids pass more easily into a
cell than those that dissolve in water. He then set forth an equation that predicted
how fast that diffusion would happen. One of the key parameters in that equation
is K which defines the lipophilicity (oil-liking nature) of the chemical. The
higher the value of K, the faster the predicted cell permeation rate. For over
a century, medicinal chemists have used this relationship to shape their studies
and clinical trials.
A team of electrochemists from the University of Warwick used a combination
of a confocal microscope and an ultramicroelectrode to study what really happens
when a chemical crosses a cell membrane. Advances in technology enabled them
to position an ultramicroelectrode incrediblely close to the membrane boundary
(roughly 20 microns away; ca. 1/3rd the thickness of a human hair) where it
was used to generate a range of acids that should be able to diffuse relatively
easily into a cell. These techniques allowed every step of the diffusion process
to be directly examined. Previous studies had not been able to observe every
step of the process and often required artificial stirring of the solutions.
The results stunned the researchers. While the acids did diffuse across a lipid
membrane, they did so at rates that were diametrically opposite to the predictions
of the Rule, i.e. the most lipophilic molecules were actually transported slowest.
The researchers studied four acids (acetic, butanoic, valeric, and hexanoic)
that had increasingly larger "acyl" (or carbon) chains. The longer
the carbon chain, the easier the chemical dissolves in lipids and, therefore,
according to Overton, the faster they should diffuse across a lipid membrane.
In fact, the University of Warwick researchers observed that for these four
acids the exact opposite is true: the easier it is for an acid to dissolve in
a lipid, the slower it is transported across the membrane.
The research team will now use their technique to examine the diffusion into
cells of a range of other chemicals. The lead researcher on the study from the
University of Warwick, Professor Patrick Unwin, said:
"This was a surprising and exciting finding. Our direct observations appear
to totally undermine a key rule that has withstood the test of time for over
a century. We will now make observations with a range of other chemicals, and
with other techniques, to further elucidate the molecular basis for our observations.
Text books will have to be rewritten to revise a rule that has been relied on
for over a century. Advanced techniques, such as the one we have developed,
should give much clearer insight into the action of a wide range of drug molecules,
which will be of significant interest to drug developers."