Nucleic acid therapeutics are based on nucleic acids or closely related chemical compounds. They include antisense oligonucleotides, aptamers and small interfering RNAs, and are typically considered in cases where specific inhibition of the function of a particular gene involved in disease is thought to be therapeutically desirable.

1. A presentation
on
Nucleic Acid Therapeutics
Tasmina Ferdous Susmi

2. Outline
 Therapeutic agent & Nucleic acid therapeutic agent.
 Types of Nucleic acid therapeutic agent.
 Mechanism of Nucleic acid therapeutic agent.
 Application of Nucleic acid therapeutic agent.
 Advantages & disadvantages of Nucleic acid therapeutic agent.
 Future aspects of Nucleic acid therapeutic agent.

3. Therapeutic agents
The term comes from the Greek therapeutikos word which
means “inclined to serve”.
Therapeutics means serving and caring for the patient in a
comprehensive manner, preventing disease as well as managing
secific problems.
Relating to therapeutics, the branch of medicine that is
concernsed specially with the treatment of disease.
Therapeutic dose of a drug is the amount needed to treat a
disease

4. Nucleic acid as therapeutic agents
Therapeutic system using nucleotide
sequences are known as nucleic acid
therapeutic agents.
The delivery of nucleic acid molecules
into cells to alter physiological
functions at the genetic level is a
powerful approach to treat a wide
range of inherited and acquired
disorders.

5. Nucleic acid as therapeutic agents
Insertion of new genetic material
into the cells of an individual can
bring intentional therapeutic
benefit
Numerous nucleic acid-based
therapy are under development.
Some of which is used to inhibit
gene expression at either the
transcriptional or post
transcriptional level.

6. Why Nucleic acid therapeutics ?
To treat human diseases, researchers must be able to safely and
effectively deliver nucleic acids to their targets.
Nucleic acids interference therapies have the potential to treat
many human diseases.
Particularly those stemming from the gastro-intestinal (GI) tract,
by transferring genetic materials to diseased cells to inhibit
abnormal gene expression.

7. Nucleic acid as therapeutic agents
Delivery vectors must protect the encapsulated material while
traveling through the GI tract, and also be safe and easy to
tolerate.
Orally delivering nucleic acid therapy is a particularly
promising option because it is simple to administer, non-
invasive, and cost-effective.
However, only a small percentage of these doses travel
successfully to the disease site. They are impeded by
anatomical and physiological barriers, including rapid
degradation at low gastric pH and enzymatic degradation in
the intestines.

8. Types of Nucleic Acid Therapeutics
RNA based therapeutics:
Antisense RNA
Ribozymes
RNA aptamers
Interfering RNAs
DNA based therapeutics:
Plasmid DNA
Antisense oligonucleotide
DNA aptamer
Deoxyribozymes
Chimeric DNA-RNA molecules

9. RNA based therapeutics
Antisense RNA
Single stranded RNA molecule
complementary to specific mRNA.
15-20 nucleotides
Technology for prevention of
expression of a gene translation level.
Bind mRNA then take the help of
RNAse H to breakdown the mRNA
selectively.

10. RNA based therapeutics
Ribozymes:
Naturally occurring catalytic molecules
that are 40-50 nucleotides in length.
Have separate catalytic and substrate
binding domains.
Important advantages of ribozymes is
that they are unlikely to evoke an immune
response in a treated animal or human.
Example: Self splicing
intron,SnRNAs,RNAse P,
Hairpin,Hammerhead.

11. RNA based therapeutics
RNAAptamer:
Single stranded nucleic acid segments, can
directly interact with protein.
The Word aptamer, derived from the latin
"aptus"-fit and greek " meros"-part.
Recognize their targets on the basis of shape
complimentary.
Binding specificity and affinity for target is
high.
Often identified using a technology called
SELEX.
Clinical study on humans with injection of
anti VEGF aptamers in eye.

12. RNA based Therapeutics
Interferring RNA:
Part of transcriptional mechanism of gene
silencing through chromatin
remodeling,inhibition of protein or direct
mRNA degradation.
Used for down regulation of disease causing
gene through RNA interference.
Double stranded RNA molecules are
cleaved by RNase lll like enzymes into single
stranded RNA(21-23 nucleotides) called
small interfering RNA.
Inhibit HIV,Hepatitis,Influenza infection.

13. DNA based therapeutics
Plasmid DNA:
High molecular weight,double stranded DNA constructs
containing transgene encode specific protein.
Used in gene therapy.
Used to correct genetic errors that produce funtionally
incompetent copies of a given protein.
Antisense oligonucleotide:
Oligonucleotide which is short single stranded segments of DNA
that upon intematization can selectively inhibit the expression of
single protein.
Form duplex with mRNA and inhibit protein biosynthesis.

14. DNA based therapeutics
DNAAptamers :
ds nucleic acid segments that can directly interact with proteins.
Interfere with molecular function of disease implicated protein or
those that participate in transcription or translation processes.
Preferred over antibodies in protein inhibition owing to their
specificity,non-immunogenecity and stability.
Deoxyribozymes:
Analogues of ribozymes with greater biological activity.
Used against cardiovascular disease,cancer.

15. Chimeric RNA-DNA molecules
Includes single mixed
oligonucleotides with ribonucleotide and
deoxyribonucleotides in a duplex
conformation.
In organization of nucleotides
chimeric oligonucleotide is important.
Protect the molecules from
exonuclease.
Shields the molecules from
degradation of RNase.
Helps in basepair correction.

16. Mechanism of Antisense RNA
therapeutics
 Single stranded RNA.
 Complementary to mRNA.
 Introduced in a cell to inhibit it's
translation machinery.
 Binds with mRNA & activates RNAse
H enzyme.
 RNA duplex is unable of translation.
 Thus stops the production or activity
of a specific protein.

17. Mechanism of Antisense RNA
therapeutics(cont.)
AUGAAA- Sense
RNA/mRNA
UACUUU- Antisense RNA

18. Mechanism of Antisense oligonucleotide
 Hybridizes with complementary
nucleotide sequence/mRNA.
 Blocks the production of specific
protein.


19. Mechanism of Ribozyme


20. Mechanism of Ribozyme

21. Mechanism of Ribozyme

22. Mechanism of Aptamer


23. Mechanism of Aptamer

24. Mechanism of RNAi
 Double stranded RNA containing
complementary sequence of target
mRNA is used.
 Dicer enzyme cuts the RNAi into
siRNA's.
 Antisesne siRNA strand
incorporated into RISC(RNA
induced silencing complex).
 RISC binds to the mRNA and
cuts it.

25. Mechanism of RNAi

27. Applications
For treatment of any human disorder caused by over production of
normal protein e.g,
Cancer
Inflammatory conditions
Viral and parasitic infections

28. Applications
• Antisense RNA
 For treatment of malignant glioma(Human brain tumor)
 For treatment of prostate carcinoma
• Antisense oligonucleotide
For treatment of cytomegalovirus infection
To prevent restenosis after angioplasty
Smooth muscle cell proliferation implicated in
atherosclerosis, hypertension, diabetes mellitus, and the
failure of coronary bypassed grafted are presumably
controlled by antisense therapeutics.
To control psoriasis(Uncontrolled epidermal growth)
For treatment of tuberculosis

29. Applications
 Correction of a mutant splice site with an antisense
oligonucleotide can be used to treat β-thalassemia.

30. Applications
• Ribozymes
To prevent viral infections
To prevent the accumulation of chemical/antibiotic
resistance
• RNA Interference (RNAi)
Gene silencing which plant and animals use to protect
against viruses.
To treat human neurodegenerative disease
spinocerebellar ataxia 1
Scientists are optimistic about using RNAi to treat other
neurological diseases such as Alzheimer’s and
Hunting’s disease.

31. Applications
DNA vaccine
Antibody genes

32. Advantages
Highly specific/selective
Functional diversity
Limited toxicity
High efficiency

33. Limitations
Susceptible to degradation by cellular nuclease enzyme.
Less stability inside cell
Need to be modified which may cause off target
Determination of appropriate concentration of nucleic acid
Viral vector may pose danger
To deliver nucleic acid in target sites is difficult

34. Present situation of nucleic acid
therapeutic agents
Nucleic acid therapeutic agents have been researched as
potential drug for diseases such as –
 Lung cancer
 Colorectal carcinoma
 Pancreatic carcinoma
 Malignant glioma
 Malignant melanoma
 Diabetes
 Amyotrophic lateral sclerosis
 Duchenne muscular dystrophy
 Spinal muscular atropy
 Asthma

35. Cancer
 To treate gliomas
 AP12009-
 A phosphorothioate antisens
oligodeoxynucleotide specific for
the mRNA of human transforming
growth factor TGF-beta2
 Experiment on 145 people
-38 anaplastic astrocytoma
patients treated with Karnofsky
-89 anaplastic astrocytoma
and glioblastoma patients treated
with AP 12009
- 12 patients were treated with
TMZ or PCV

36. Cytomegalovirus retinitis
 Also known as CMV retinitis
 Is an inflammation of the retina
of the eye that can lead to
blindness
 Caused by a type of herpes virus
called Cytomegalovirus
 Treated with
 -Fomivirsen (Vitravene )
 Approved by Food and Drug
Administration

37. Familial hypercholesterolemia
 Known as homozygous familial
hypercholesterolemia, or HoFH
 Causes extremely high cholesterol level and
heart attacks
 Treated with Mipomersen (kynamro)
 Approved by Food and Drug Administration

38. Hemorrhagic fever
 In 2006 – 75% recovery rate
 Ebola hemorrhagic fever virus
 Treated with Morpholino
 Developed by sarepta therapeutics
HIV/AIDS
 Starting 2004
 Virus escape mutants
 pVRX494 and pVRX496 plasmid –derived from
pUC18 backbone

39. Spinal muscular atrophy
 In 2004
 Antisense oligonucleotide –(nusinersen),
developed by Ionis Pharmaceuticals
 FDA approval 2016
Duchenne muscular dystrophy
 A severe types of muscular dystrophy
 X-linked recessive
 Mutation of the gene for the protein dystrophin
 Treated with eteplirsen(a Morpholino oligo)
approved by FDA September 2016

40. Hypertriglyceridemia
 December 2016
 Treated by volanesorsen
 Volanesorsen is in phase 3 clinical trials

41. Future aspects
Antisense RNA and oligonucleotides :
 Lung cancer,
 Colorectal carcinoma,
 Pancreatic carcinoma,
 Malignant melanoma,
 Diabetes,
 Asthma
 Arthritis

42. Future aspects
Antisense RNA and oligonucleotides :
 Lung cancer,
 Colorectal carcinoma,
 Pancreatic carcinoma,
 Malignant melanoma,
 Diabetes,
 Asthma
 Arthritis

43. Future aspects
Aptamers :
 Therapeutic application of aptamers in lymphoma
 Aptamers “small bomb” mediates delivery of drug to cancer cells
 Potential next generation of diagnostic tools
 New drug development
 Bio-imaging
 Drug discovery

44. Future aspects
Interfering RNAs:
 Used in treating cancer
 Defense against infection by viruses
 Suppress the transcription of viral genome
 Used to treat bacterial diseases
 Potential for treating neurodegenerative disease(Parkinson's
disease and Polyglutamine disease)

45. Thank's to everybody for
your kind attention