Abstract

Target Identification and Validation investigations largely
depend on gene silencing studies as it remains the most widely used approach to
study gene function in order to establish therapeutic relevance. Standard gene
silencing methods using RNAi or CRISPR technologies are efficient and useful. However, both depend on recruiting
cellular machineries, the RISC complex for RNAi and the DNA repair non-homologous
end joining (NHEJ) for CRISPR. Obviously these two machineries are in the cell for
reasons not for the experimental scientists’ “exploitations”. In normal cells,
the RNAi’s RISC complex is used by endogenous microRNA pathway, as it is an
important layer of gene regulators. Whereas, the CRISPR’s NHEJ is used in DNA repair, as
it is an important phase of the cell cycle. Consequently, some sort of
disturbance to the normal cellular activities and behaviour
may occur as a result of recruiting RISC or NHEJ,
which may lead to abnormal undesired outcome. This is in addition to the well-documented
off-target effects of both RNAi and CRISPR that have been widely addressed in
the literature.

Thus, it would be
useful to consider the use of an alternative cellular independent gene
silencing mechanism, such as the catalytic cleavage of mRNA using the catalytic
DNAzymes.

DNAzymes represent a class of
catalytic nucleic acids that have the ability to cleave RNA substrate in an
enzymatic fashion without any requirements of any cellular machineries. DNAzymes
are currently used as therapeutic agents in the clinic after successful
clinical trials^1-2.  Discovered long before the RNAi, DNAzymes
used then for gene silencing but with limited popularity due to their optimisation
requirement and cellular instability. We previously reported novel ways of
addressing these two limitation particularly by introducing the Hairpin
DNAzymes.

Hairpin DNAzyme (hpDz)³ is a modified DNAzyme that
contains stem loop at the binding arms. hpDz showed stability against
endogenous cellular nucleases and also efficient gene silencing.

Today, in the relevance
of Target Identification and Validation, we believe that considering an alternative
gene silencing tool that is independent of cellular machineries may contribute
to a better validation for therapeutic targets. Moreover, hpDz may be also
considered as therapeutic agents.

We believe that in the light of the growing concerns of RNAi
and CRISPR off-target effects, there is a need for an alternative gene silencing method that is
independent of using any cellular machineries and has no off-target effects.

OMMTech is specialised in
Target Validation as a CRO and offers hpDz technologies as one of its
services at two formats:

a)      
Hairpin
DNAzymes Kits – custom-made pre-optimised hpDzs tailored to clients’
requested genes. The kit comes with a 6-month free consultation to assist
clients’ experimental work.

b)     
Hairpin
DNAzymes Projects - We will compare your RNAi or CRISPR gene silencing data
with DNAzymes technology in order to reassure reproducibility and
target-phenotype association using different silencing technologies.