1) Genome editing-based therapies are a promising alternative to highly-active antiretroviral therapy (HAART) for treating HIV infection. These therapies aim to produce immune cells resistant to HIV using tools like zinc finger nucleases, TALENs, and CRISPR/Cas9. 2) The major strategy involves modifying cells like hematopoietic stem cells and T cells in vitro before reinfusing them into patients. Recent genome editing therapies have targeted HIV genes like CCR5, CRX4, and the HIV provirus. 3) While genome editing therapies for HIV have made progress and entered clinical trials, concerns remain around their safety, choice of targets as HIV can switch receptors

1. GENOME EDITING-BASED (GE-Based) HIV
THERAPIES
BY
ICHIPI-IFUKOR, Patrick Chukwuyenum
PG/13/14/223056
A SEMINAR PRESENTED
IN
The Department of Biochemistry, Faculty of
Science
Delta State University, Abraka

2. INTRODUCTION
Acquired immunodeficiency syndrome (AIDS) is a
serious infectious disease characterized by the
systemic collapse of the immune system caused by a
retrovirus designated HIV-1. The pathogen was
identified 2 years after the first report of the disease in
1981(Gu, et al., 2014).
The entry of human immunodeficiency virus (HIV) into
cells is critically dependent on the sequential
interaction of the viral envelope with two cell-surface
receptors, the CD4 glycoprotein and a 7-
transmembrane-domain chemokine receptor such as
CCR5 or CXCR4.
The HIV-1 genome also consists of nine genes (Gag,
Pol, Env, Vif, Vpr, Vpu, Tat, Rev, and Nef). A LTR

3. RECEPTOR INTERACTIONS INVOLVED IN HIV ENTRY.
Figure 1: Receptor interactions involved in HIV entry.
Clapham, and McKnight , 2001
(A) HIV virion binds CD4.
(B) CD4 binding induces
conformation changes in
gp120 that result in the
movement of the variable
loops and exposure of the
co-receptor binding site.
Flexible regions in CD4
between domains 2 and 3
as well as between
domain 4 and the
membrane allow
orientation of the co-
receptor binding site for
co-receptor binding.

4. Figure 2: HIV infection, replication cycle and the HIV genome. (A) Co-receptor
usage shift during different stages of infection progression. (B) The HIV
replication cycle. (C) The HIV-1 genome.
Source: Gu, 2015

5. WHYGENOME EDITING-BASED HIV THERAPIES
 The only effective therapy used in clinical treatment
for HIV infection is highly-active antiretroviral therapy
(HAART), which is a combination of antiviral drugs
targeting different steps in the virus life cycle.
Theoretically, each step of HIV replication could be
targeted for anti-HIV therapy. However, the HIV-1
genome consists of nine genes, which are also
important targets for anti-HIV research
Developing molecular therapeutics for these targets
could be viable, but GE-based therapies stand out as
a possible alternative to HAART for the treatment of
HIV infection.

6. BASIC STEPS APPLIED IN GE-BASED
THERAPIES
The major strategy for GE-based HIV therapies is to
produce engineered immune cells that are resistant to HIV
infection or replication. The possible cell types for
modification range from hematopoietic stem cells to
dendritic cells and CD4+ T cells (Rossetti, et al., 2012).
The most frequently used method consists of two steps:
modifying the cells in vitro then reinfusing the modified
cells into patients (Figure 1) (Voit, et al., 2013)

7. Figure 3: Basic Steps involved in Genome Editing Based HIV Therapies
Source: Rossetti, et al., 2012; Voit, et al., 2013
A B

8. Table 1: New Genome Editing Technologies/Tools
Nuclease Origin Delivery
Tools
Advantages Disadvantages
Zinc Finger
Nucleases
(ZFN)
Eukaryotes AV, AAV,
LV
Compatible with various viral
vectors; each ZF module (30 amino
acids) recognizes three sequence-
specific nucleotides; widely used in
various cell types and multiple
organisms including humans
Design of ZF arrays
for sequence
recognition is
required; off-target
effects
Transcripti
on
activator-
like
effector
nucleases(
TALEN)
Xanthomo
nas
AV Each TALE repeat (33–35 amino
acids) recognizes one nucleotide;
widely used in various cell types
and multiple organisms including
humans
Design of TALE arrays
for sequence
recognition is
required; the binding
site starts with a T
base; larger size than
ZFN; off-target effects
Clustered
Regularly
Interspace
d Short
Palindromi
c Repeats
(CRISPR/
CAS )
Bacteria Plasmids RNA-guided DNA endonuclease;
targets multiple sites
simultaneously to produce large
gene fragment deletions; much
simpler design (a short RNA for the
gene of interest); potential
application in various hosts
including humans
The target sequence
needs to be preceded
by PAM; off-target
effects
Source: Gu, (2015)

9. Table 2: Summary of Recent GE-based HIV therapies
Target gene Editing tool Target Cell Reference
CCR5 ZFN Autologous CD4+
T cells (clinical
trial)
ClinicalTrials.gov
(2014)
CRX4 ZFN Infected T cells Wilen, et al., 2011
CCR5 ZFN/TALEN 293 T cells
Provirus (LTR) ZFN Infected T cells Dats and Sagal,
2014
Provirus (LTR) CRISPR/Cas9 Infected T Cells Ebina, et al., 2013
Provirus CRISPR/Cas9 Microglial,
promonocytic, and
T cells
Hu, et al., 2014
LEDGF/p75 TALEN 293 T cells, Jurkat
cells
Christ and
Debyser, 2012
Source: Gu, (2015)

10. Concerns for GE Based Therapies
Although great progress has been achieved in
GE based-therapies, and many have proceeded
into clinical trials, several issues give cause for
concern.
Safety is a fundamental concern in GE-based
therapies.
 The choice of target for these therapies: CCR5
serves as the predominant co-receptor in the
early stage of HIV infection, and CXCR4 plays a
key role as co-receptor when stable infection is
established. HIV co-receptor usage can switch
between CCR5 and CXCR4 resulting in failure of

11. The difficulties in finding HLA-matched donors
with homozygosity for the CCR5 d32 deletion, as
well as the problems in the transplant process,
undercut the feasibility of deploying this therapy.
A technical concern for GE tools is off-target
cleavage events, which can lead to unintended
disruption of physiologically important genes, and
can have unpredictable consequences.

12. Conclusion
Although, the current GE-based HIV therapies are
mainly based on reinfusion of modified CD4+ T
lymphocytes or CD34+ stem cells, scientists must
develop cheaper and more affordable technologies that
can be easily accessible. While most economies must
remain determined to winning the war against HIV/AIDS
through funding.

13. THANK
YOU FOR
LISTENING