RNA interference (RNAi) is a powerful strategy for suppressing gene
expression in a sequence-specific manner. This strategy offers new potential
opportunities for treating various diseases by addressing otherwise
'undruggable' targets. Currently, the most promising type of RNAi-based
advancing therapeutics in preclinical and clinical trials is incorporating small
interfering RNAs fragments (siRNAs), synthetic 21-23 base pairs double-stranded
RNA molecules, into the cell cytoplasm1,2.
RNA interference (RNAi) is a natural
process that cells use to turn down, or silence, the activity of specific genes.
Discovered in 1998, RNAi has taken the biomedical community by storm.
Researchers quickly capitalized on the discovery and developed RNAi into a
powerful research tool that is now used in thousands of labs worldwide.
Gene silencing is the interruption or suppression of the
expression of a gene at transcriptional or translational levels. Scientists have
been working on strategies to selectively turn off specific genes in diseased
tissues for the past thirty years.
Leukocytes are a type of immune cell. Most leukocytes are
made in the bone marrow and are found in the blood and lymph tissue. Leukocytes
help the body fight infections and other diseases. Granulocytes, monocytes, and
lymphocytes are leukocytes. Also called WBC and white blood cell.
Cytoplasm is the fluid inside a cell
but outside the cell's nucleus. Most chemical reactions in a cell take place in
Gene expression is process by which a
gene gets turned on in a cell to make RNA and proteins. Gene expression may be
measured by looking at the RNA, or the protein made from the RNA, or what the
protein does in a cell.
Despite advantages such as eliminating clinical safety concerns associated
with viral vectors and a lesser interruption to endogenous gene regulation
machineries, translation of siRNAs into clinical practice faces some major
hurdles, like low efficacy of crossing the plasma membrane and entering the
cytoplasm, stimulation of the immune system that often causes global suppression
of gene expression, rapid renal clearance and degradation by RNases. Therefore,
generating nanocarriers for targeted delivery of siRNAs is necessary. Devising
such systems enforces dealing with the challenges of developing fully degradable
particles targeted to the appropriate cell-type, evading the stimulation of the
immune system, and utilizing cellular mechanisms for internalization and
releasing the siRNAs into the cell cytoplasm2,3.
Unlike systemic delivery to solid tumors and the liver, systemic delivery to
leukocytes (immune cells), due to their resistance to conventional transfection
methods and to their dispersing in the body, remains challenging4. We have developed antibody-protamine (a positively
charged protein) fusion proteins directed to the lymphocyte function associated
antigen-1 (LFA-1) integrin, a cell surface adhesion molecule that is expressed
in all leukocytes' subtypes. Those fusion proteins selectively delivered siRNAs
into leukocytes, both in vitro and in vivo. Furthermore, by targeting these
fusion proteins to the high affinity conformation of LFA-1 that characterizes
activated lymphocytes, we demonstrated selective gene silencing, which unlike
most immunosuppressive therapies, could provide a way to overcome the unwanted
immune stimulation without global immunosuppressive effects on bystander immune
order to increase payload and achieve robust targeted gene silencing, we have
generated integrin-targeted stabilized nanoparticles (I-tsNP), which have been
covalently coated with anti-B7 integrin (highly expressed in gut mononuclear
leukocytes) antibody, and demonstrated that those particles can selectively
deliver siRNAs to leukocytes involved in gut inflammation (see illustration).
Made from natural biomaterials, these nanoparticles offer a safe platform for
siRNAs delivery, avoiding cytokine induction and liver damage6. Using this system, we identified cyclin D1, a regulator
protein of the entry into, and the progression throughout the cell cycle7, as a potential new target for treating
|Integrin-targeted stabilized nanoparticles (I-tsNP) . The
particles have been developed as ~80nm liposomes, formed from natural
phospholipids, hence avoiding the potential toxicity of cationic lipids and
polymers. Hyaluronan (HA), a naturally accruing glycosaminoglycan, was attached
to the surface of the liposomes, stabilizing the particles during siRNA
entrapment and systemic circulation in vivo. Then, a monoclonal antibody against
the integrin was attached to HA. The particles were loaded with siRNAs condensed
with protamine while maintaining their
Utilizing siRNAs to manipulate gene expression in leukocytes holds great
promise for the drug discovery field, as well as for facilitating the
development of new therapies platforms for leukocytes implicated diseases such
as inflammation, blood cancers, and leukocytes-tropic viral infections.
Additionally, siRNA delivery to leukocytes could serve as a powerful tool for
understanding leukocytes' biology. Moreover, since one can easily change the
payloads inside the nanoparticles (by using different sequences of siRNAs, or
other drugs) or the targeting agent (by replacing the antibody or the ligand
decorating the nanoparticle's surface), it is reasonable that this platform
might be applicable to other types of diseases outside the hematopoietic
1. Sledz CA and Williams BR. RNA interference in biology and
disease. Blood, 106, 787-794 (2005).
2. de Fougerolles A,
Vornlocher HP, Maraganore J, Lieberman J. Interfering with disease: a progress
report on siRNA-based therapeutics. Nature reviews, 6(6), 443-453 (2007).
3. Peer D, Karp JM, Hong S, Farokhzad OC, Margalit R, Langer R.
Nanocarriers as an emerging platform for cancer therapy. Nature nanotechnology,
2(12), 751-760 (2007).
4. Goffinet C, Keppler OT. Efficient
nonviral gene delivery into primary lymphocytes from rats and mice. Faseb J,
20(3), 500-502 (2006).
5. Peer D, Zhu P, Carman CV, Lieberman
J, Shimaoka M. Selective gene silencing in activated leukocytes by targeting
siRNAs to the integrin lymphocyte function-associated antigen-1. Proceedings of
the National Academy of Sciences of the United States of America, 104(10),
6. Peer D, Park EJ, Morishita Y, Carman CV,
Shimaoka M. Systemic leukocyte-directed siRNA delivery revealing cyclin D1 as an
anti-inflammatory target. Science, 319(5863), 627-630 (2008).
7. Stacey DW. Cyclin D1 serves as a cell cycle regulatory switch in
actively proliferating cells. Curr Opin Cell Biol, 15, 158-163 (2003).
Copyright AZoNano.com, Dr. Dan Peer (Tel Aviv