**5.5. Genome editing**

agents. As most of the shocking agents are related to anti-cancer drugs, they are likely to have a significant adverse effect profile. The activators of the NF-κB pathway are notorious for triggering a plethora of cytokines and can cause a fatal cytokine storm if administered at higher concentrations. Another important drawback is that, even if an efficient shocking agent is identified, killing of the actively replicating virions depends on anti-retroviral drugs which are expected to lose their efficacy in the future due to the evolution and dissemination of resistant viral population. Nevertheless, this strategy has immense potential in eradicating

Apart from the trial of different compounds, techniques of gene therapy have also been tried out to activate viral reservoirs with limited success. Herpes virus and lentiviral vectors loaded with genes coding for key viral proteins involved in replication were found to induce repli‐ cation and release of the provirus from the latent CD4 T-cells [131]. Problems with *in vivo* administration, lack of specificity in targeting the cellular reservoir, low frequency of recom‐ bination are the various setbacks which have withheld the progress of this technology in

Another interesting finding pertaining to reservoir eradication is the property of the gold complex drug 'auranofin' to selectively destroy the retroviral cellular reservoir. Although developed and used as an anti-rheumatic agent, the unique 'anti-memory T-cell effect' of auranofin has kindled interest in its possible role against HIV. Auranofin exerts its cytocidal action by inducing intracellular oxidative stress. The memory T-cells with low antioxidant defenses are highly vulnerable to the oxidative stress induced by auranofin and perish along with the integrated provirus. Combination of auranofin and buthionine sulfoximine, an inhibitor of glutathione synthesis is found to act synergistically by causing further imbalance

Auranofin has shown promising results in studies utilizing the simian AIDS model. Simian immunodeficiency virus (SIV) infected macaques, treated with a combination regimen of auranofin, buthionine and HAART have not only shown a prolonged post treatment drug free control of viremia but also developed enhanced cell mediated immunity with SIV specific cytotoxic CD8 T-cells following treatment suspension [134, 135]. However, well designed human clinical trials are required to know more about this anti-HIV reservoir compound

With the mechanism exactly opposite to the shock and kill strategy, this strategy aims at achieving viremic control by keeping the viral reservoirs continually in the inactive state. As mentioned under the shock and kill strategy, concurrent activation of the gatekeeper kinase and NF-κB favours reactivation of the provirus. The compound Jun N-terminal protein kinase inhibitor-5 which is a potent inhibitor of the gatekeeper kinase strongly prevents viralreactiva‐ tion even upon strong stimulation of the NF-κB pathway under *in vitro* conditions. Inhibitors

viral reservoirs if the various setbacks are addressed over time [130].

226 Trends in Basic and Therapeutic Options in HIV Infection - Towards a Functional Cure

reservoir elimination [132].

in the redox pathways [133].

**5.4. Repression of latency**

**5.3. Selective cytolysis without viral reactivation**

before its flamboyance could be translated to practicality.

Genome editing techniques confer the possibility of excising out specific genetic sequences from the whole genome. This novel concept is currently being used for research involving in eukaryotic genome manipulation to produce gene knockout animals. The ability of this technology to specifically excise the integrated provirus from the genome of the latent reservoir cells is being evaluated.

Three different genome editing systems are currently available such as the zinc finger nucle‐ ase (ZFN) system, the transcription activator like-effector nucleases (TALEN) system and the clustered regularly interspaced short palindromic repeat (CRISPR) with CRISPR-associated protein-9 (Cas9) known as the CRISPR/Cas9 system (Table-5). Of the three systems, the ZFN and the CRISPR/Cas9 system are being tried in the genome editing of the latent reservoir cells. *In vitro* studies employing the CRISPR/Cas9 system have demonstrated the ability of the technique to remove the HIV internal genes and suppress proviral reactivation in T cells [138]. Alternatively,studiesemployingtheZFNsystemhavedemonstratedtheabilityofthetechnique to excise the full length HIV proviral DNA from the infected human T cell genome [139].

This advanced technology also has its own drawbacks. The efficacy of genome editing strategies observed with different *in vitro* studies is about 30% though research work is still ongoing, addressing the challenges and improving on the efficacy. Suitable transport systems are yet to be developed for *in vivo* delivery of the editing machinery to the viral reservoir cells. All genetic editing systems have a certain proportion of 'off-target activity' where they excise out genes unrelated to the targeted site. As this can result in undesired gene modification events, the genome editing systems must be made more specific before its utilization for anti-HIV treatment [140].
