Ashok Chauhan talks with
ScienceWatch.com and answers a few questions about
this month's Fast Breaking Paper in the field of
Pharmacology & Toxicology.
Article Title: The taming of the cell penetrating
domain ofthe HIV Tat: Myths and realities
Authors: Chauhan,
A;Tikoo, A;Kapur, AK;Singh, M
Journal: J CONTROL RELEASE
Volume: 117
Issue: 2
Page: 148-162
Year: FEB 12 2007
* Johns Hopkins Univ, Dept Neurol, Richard Johnson Div
Neuroimmunol & Neurol Infect, 600 N Wolfe St,509
Pathol, Baltimore, MD 21287 USA.
* Johns Hopkins Univ, Dept Neurol, Richard Johnson Div
Neuroimmunol & Neurol Infect, Baltimore, MD 21287
USA.
(addresses have been truncated)
Why do you think your paper is highly
cited?
Our study has reported novel findings and highlights ingenious ways to
improve the delivery of small therapeutic molecules using cell-penetrating
peptides.
Does it describe a new discovery, methodology or
synthesis of knowledge?
We have systematically described our findings and the possible
repercussions of the previous studies which rely heavily on pleiotropic
effects of protein transduction domains (PTD) or Tat protein (the
transactivator protein from lentiviruses,
notably HIV) which may not necessarily be true. We have
described here new ways which may help in the development of newly
potent gene, protein, and drug-delivery vehicles for effective
biodistribution.
Would you summarize the significance of your paper in
laymen's terms?
We have elucidated several new properties of cell penetrating peptides
(PTD) as a protein delivery and gene delivery vehicle which may have the
potential for the development of novel therapeutics. We have described a
stepwise strategy which will help in producing recombinant proteins for
successful biodistribution.
"Our aim is
to look for an aggressive way to
curtail
HIV for the long term
and for this we plan to impede the
HIV life cycle at different steps,
using our peptide in different
forms.."
In this manuscript, we have elucidated the possible mechanism of fusion
protein entry into cells and resistance encountered by fusion proteins
after entry. We have described ways to overcome the intracellular
resistance and hence enable effective delivery and biodistribution. We have
further described a way to predict the nature of fusion protein after
fusion with PTD sequence and its ultimate fate (distribution) inside the
cell.
We have highlighted shortcomings of this peptide (PTD) in general as a weak
delivery vehicle, owing to its inefficient entry on fusion with different
proteins. We infer that this peptide will be useful in delivering proteins,
RNA, and DNA sequences; however, not in the native form. PTD's potential
can profoundly be enhanced by fine-tuning of peptide sequence using
available knowledge and cutting-edge technology.
How did you become involved in this research, and were
there any problems along the way?
I started doing research on HIV-Tat protein and PTD in 1996 while working
as a visiting scientist in the Departments of Neuroscience and Microbiology
and Tumor Biology, at the Karolinska Institute in Sweden, and then I
continued here in the United States at Johns Hopkins University in
Baltimore. I am a virologist trained in RNA viruses and my research for the
last 12 years has focused on HIV.
In the HIV life cycle, early Tat-protein has been described as pleiotropic
in function apart from its primary transactivation function in HIV
replication. I was working on HIV gene therapy to inhibit Tat protein and
cripple the HIV life cycle. Tat protein is a mastermind of the HIV life
cycle and its inhibition will curtail HIV-multiplication.
Tat protein has a unique property of entering other cells when added
externally. This property of Tat is localized to a short stretch of
sequence (PTD) in the middle of the protein. Since PTD is responsible for
entry into the cell, I was specifically interested in delivering
fusion-proteins into neurons. After complex designing and cumbersome
experimentation with repeated failures, I finally succeeded in introducing
fusion proteins into neurons and other cells. However, protein entry was
not as impressive as was anticipated from earlier studies.
On further modifications in design and experiments, we came to the
conclusion which was published in the Journal of Controlled
Release in 2007. In science, when you are looking for something, you
normally do not find precisely what you are looking for, instead you often
discover something else. It is difficult to publish new concepts because
people normally do not accept new ideas quickly. We encountered stiff
resistance to publishing our findings; however, some people found our
findings of interest and recommended that they be published.
Where do you see your research leading in the
future?
We are headed towards the use of PTD or small cell-penetrating peptides and
their permutations in delivering small therapeutic molecules such as siRNA
and antisense. Small interfering RNA (siRNAs) are small synthetic
inhibitors (gene silencers) designed to target specific viral or
other genes, in our case HIV. We are at the final stage of concluding
our investigations on the novel applicability of peptide-siRNA in
HIV-inhibition. We believe that for safe drug and gene therapy in the
future, non-viral delivery methods will be the ultimate choice and will
be helpful in site-specific delivery of RNA and DNA sequences to correct
disease states.
However, this kind of research has not yet picked up pace with other
delivery systems. Indeed, we are fine-tuning these peptide sequences to
give them target specificity and to have more potent and specific effects.
Target-specific delivery will be our ultimate goal.
Do you foresee any social or political implications for
your research?
Our aim is to look for an aggressive way to curtail HIV for the long term
and for this we plan to impede the HIV life cycle at different steps, using
our peptide in different forms. HIV is a very complex virus which affects
millions of people around the globe. The disease is possible to treat,
however, and difficult but not impossible to eradicate. Presently, the HIV
life cycle is known somewhat, but not quite enough to control its activity
completely.
Hence we are investigating the HIV life cycle in order to understand its
weaknesses and to target those steps to give a death blow to the virus. Our
research will have social and political implications since HIV, in
particular, is a disease without borders. We also predict that our research
will have implications in the rational designing of vaccines, potent
therapeutic drugs, and in the novel development of gene delivery vehicles
which will have implications for other chronic diseases such as
neurodegeneration.
Ashok Chauhan, Ph.D.
Associate Professor
Department of Pathology, Microbiology, and Immunology
Adjunct Associate Professor
Department of Physiology, Pharmacology and Neuroscience
University of South Carolina
Columbia, SC, USA
Keywords: protein transduction domain (PTD), HIV Tat, protein delivery,
cell penetrating peptides, HIV gene therapy, Small interfering RNA
(siRNAs), peptide-siRNA, HIV-inhibition, HIV life cycle.