CRISPR-mediated gene activation, calledCRISPRa, uses dCas9 fusion proteins to recruit transcription activators. Afusion of dCas9 with the ?-subunit of the E. coli Pol allowed assembly of theholoenzyme at a target promoter for gene activation in E.
coli 20. There are currently limitedreports on CRISPRa in bacteria, and more work is needed to achieve robust andconsistent gene activation in bacteria 5. CRISPR–dCas9 can target several genessimultaneously by using multiple sgRNAs. Recently, a method for simultaneousrepression and activation of genes was established using scaffold RNAs (scRNAs)21.ACUTE MYELOIDLEUKEMIA (AML)22 identified additional therapeutic targets inacute myeloid leukemia (AML), they optimized a genome-wide clustered regularlyinterspaced short palindromic repeats (CRISPR) screening platform and use it toidentify genetic vulnerabilities in AML cells. They identified 492 AML-specificcell-essential genes, including several established therapeutic targets such asDOT1L, BCL2, and MEN1, and many other genes including clinically actionablecandidates, validated selected genes using genetic and pharmacological inhibition,and chose KAT2A as a candidate for downstream study. KAT2A inhibitiondemonstrated anti-AML activity by inducing myeloid differentiation andapoptosis, and suppressed the growth of primary human AMLs of diverse genotypeswhile sparing normal hemopoietic stem-progenitor cells, results proposed thatKAT2A inhibition should be investigated as a therapeutic strategy in AML andprovide a large number of genetic vulnerabilities of this leukemia that can bepursued in downstream studies.
23 Using the IDH2 R140Q mutation as a model,presented a new effective methodology here using the RNA guided clusteredregularly interspaced short palindromic repeats (CRISPR)-Cas9 system toreproduce or remove AML associated mutations in or from human leukemic cells,respectively, via introduction of a DNA template at the endogenous gene locusvia homologous recombination. Our technology represents a precise way for AMLmodeling to gain insights into AML development and progression and provides abasisforfuture therapeutic approaches. Isocitratedehydrogenases (IDHs) are digestive enzymes that catalyse the oxidativedecarboxylation of isocitrate, producing alpha-ketoglutarate (a-ketoglutarate)and CO2. Mutations in IDH1/2 genes occur frequently in AML patients, and IDH2R140Q has been shown to be the most frequent IDH mutation in AML 24. IDH2 R140Q hasbeen identified as a key driver mutation in a transgenic mouse model,supporting its relevance as a therapeutic target for the treatment of human AML25.Despite intenseresearch into acute myeloid leukemia (AML) during past decades, the majority ofAML patients still die from their disease. Thus, there is a high need for newAML therapies.
AML is a heterogeneous disease harboring a multitude of geneticand epigeneticchanges,and it is highly likely that the various AML subtypes require differenttargeted therapeutic approaches 23.