Treatment of ATTR by CRISPR/CAS9 Through the Gene Editing
Treatment of ATTR by CRISPR/CAS9 Through the Gene Editing
kamran vosoo,1,*Ali kowsari,2Zeynab Vosoo,3
1. PNU International Center Island 2. Tehran University of Social Welfare and Rehabilitation Sciences 3. Gorgan University of Agricultural Sciences and Natural Resources
Introduction: transthyretin amyloidosis (ATTR) is a fatal disease which is caused by the deposition of amyloid fibrils that is included of misfolded protein in different tissues and organs. additionally, TTR, a major amyloid-genic protein, is primarily produced in the live but also in other tissues which are included choroid plexuses of the brain(1), retinal pigment epithelial (RPE) cells of the eye, and α-cells of pancreatic islets. furthermore, TTR is predominantly synthesized by hepatocytes and circulates as a homotetrameric complicated that functions as a transporter for thyroxine and vitamin A . There are 2 distinct types of ATTR: hereditary or mutated (mt-ATTR) and wild-type (wt-ATTR0). Deposition of wild-type (wt) ATTR generally happens in older patients, giving rise to wt-ATTR amyloidosis, formerly known as senile systemic amyloidosis(2). Mt-ATTR is a rare autosomal dominant condition caused by mutations in the TTR gene with considerable heterogeneity in disease presentation; Amyloid formation in ATTR is thought to happen when disconnected transthyretin (TTR) monomers misfold and collect into amyloid fibrils, with amyloid-genic mutation in the TTR gene providing the dissociation of the tetramer into monomers. Roughly, about 100 disease-causing TTR gene mutations have been reported; some scientists support the idea that to be related with particular phenotypes, however, considerable variability exists among patients. Recently, the clustered regularly interspaced short palindromic repeats (CRISPR)-related protein 9 (Cas9) system is renowned as one of the most revolutionary biological instrument. This method includes two components: the first section is that a nuclease protein Cas9 that binds to DNA and initiates double-strand breaks (DSBs), and the second one is a very shot single guide RNA (sgRNA) that directs the Cas9 nuclease to the aimed genomic locus(3). according to The research treatment agent used, for treating ATTR, it consists of NTLA-2001, is basically related to the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and associated Cas9 endonuclease (CRISPR-Cas9) system, ‘molecular scissors’ that can delete a specific disease-causing mutation at an exact location in the DNA, therefore gene function will be modified. what is more, in recent years, it has been discovered that CRISPR-Cas9 has been used to alter cells outside the body in patients with sickle cell anemia and beta-thalassemia(4).
Methods: In 1987, Ishino et al. 30 noticed in Escherichia coli, the presence of a
cluster of repetitive DNA sequences separated by variable spacer
regions. Later, Mojica et al identified identical type of repeated
sequences in numerous bacteria and archaea and named them
Clustered Regularly Interspaced Palindromic Repeats or CRISPR. 31
Interestingly, the biggest breakthrough came in 2005 when the same
group realized that these spacer sequences were from unknown ori-
gin. 32-34 Together with the observation that many CRISPR-associated
(Cas) genes encode proteins with putative nuclease and helicase do-
mains, it was postulated that CRISPR may constitute an adaptive
immunity system 33-36 by using RNAs as memory signatures of previ-
ous infections.CRISPR mechanisms are very diverse but can be mainly classified
into two distinct classes, class 1 and class 2, depending on the orga-
nization of the effector protein complex. Class 1 comprehend three
different types I, III and IV that are further subdivided into 15 sub-
types. Distinct from class 1, that is characterized by the presence of
a multi-protein effector complex, class 2 is defined by a single-pro-
tein effector module. This class is divided into types II, V and VI. 43
The other CRISPR systems have been extensively reviewed else-
where. 44,45 In CRISPR type II, DNA from viruses or plasmids of pre-
vious infections is cut into small pieces and integrated into a CRISPR
locus amongst short repetitive sequences (30-40 bp) separated by
equally short spacer sequence.There are different approaches to generate isogenic disease mod-
els in iPSCs using the CRISPR/Cas9 system.
Results: This first-ever human trial with the investigational CRISPR/Cas9-based in vivo gene
editing therapy NTLA-2001 showed a significant and consistent reduction in serum TTR
protein levels after a single admission and was generally well tolerated, representing a
potential new option for the treatment and improvement of prognosis of cardiac ATTR
amyloidosis. Continued monitoring
of whether knockout of the TTR gene results in sustained TTR reduction over the long
term is required. Evaluation of the potential effects of markedly reduced TTR levels
on patients’ clinical outcomes, with a focus on functional capacity, quality of life, andmortality benefits are essential.
Conclusion: The CRISPR/Cas9 RNA-guided DNA endonuclease system is a ver-
satile technology that has rapidly transformed genome editing andbasic science research. The development of improved CRISPR/
Cas9 tools with high degree of DNA specificity, increased selectiv-
ity and low level of by-products made this technology accessible to
researchers worldwide to study human diseases. For example, it is
now feasible to generate in vivo animal models of specific diseases in
a few weeks. It is now possible to envision the treatment of genetic
diseases in the near future using this technology. In fact, several clin-
ical trials using CRISPR/Cas9 approach to treat human genetic dis-
eases are underway.