International Journal of Molecular Medical Science, 2024, Vol.14, No.3, 193-202 http://medscipublisher.com/index.php/ijmms 196 correct the mutation in the HBB gene responsible for SCA. For instance, CRISPR/Cas9 has been used to correct the sickle cell mutation in human HSCs, which maintained the edits in vivo and produced enough normal hemoglobin to potentially benefit patients with SCA (Zeng et al., 2020). Additionally, base editing has shown potential in converting the SCD allele into a non-pathogenic variant, resulting in significant reduction of sickling in erythroid progeny (Zeng et al., 2020; Newby et al., 2021). These approaches have demonstrated the feasibility of achieving therapeutic-level gene correction in HSCs, which can be engrafted and maintained over time in animal models (Zeng et al., 2020; Uchida et al., 2021). 4.2 Animal models of sickle cell anemia Animal models, particularly mice, play a crucial role in the preclinical evaluation of gene editing strategies for SCA. Humanized sickle mouse models, which carry the human sickle hemoglobin gene, have been extensively used to study the efficacy of gene editing techniques. For example, a study using a humanized globin-cluster SCD mouse model demonstrated that CRISPR/Cas9-mediated HBB correction in HSCs resulted in stable hemoglobin-A production and normalized red blood cell features following autologous transplantation (Bak et al., 2018). These models allow researchers to assess the long-term effects of gene editing and the potential for clinical translation. 4.3 Successes in preclinical trials Preclinical trials have shown promising results in correcting the HBB mutation in animal models. For instance, CRISPR/Cas9-mediated gene correction in SCD CD34+ cells achieved therapeutic-level gene correction at both DNA and protein levels, with corrected cells contributing to normal hemoglobin production in xenograft mouse models (Figure 2) (Uchida et al., 2021). Similarly, base editing of HSCs from SCD patients resulted in high-frequency conversion of the sickle allele to a non-pathogenic variant, significantly reducing hypoxia-induced sickling in mice (Bak et al., 2018). Figure 2 Engraftment of gene-corrected SCD CD34+ cells in xenograft mice (Adopted from Uchida et al., 2021) Image caption: This figure depicts the process and results of engrafting gene-corrected SCD (Sickle Cell Disease) CD34+ cells in xenograft mice (Adopted from Uchida et al., 2021) Gene editing has also led to improvements in phenotype and survival rates in animal models of SCA. Studies have shown that gene-corrected HSCs can ameliorate the sickle phenotype, resulting in near-normal hematological parameters and reduced organ pathology. For example, lentiviral delivery of a human gamma-globin gene in a humanized sickle mouse model corrected the sickle phenotype and improved survival rates (Zeng et al., 2020). Additionally, CRISPR/Cas9-edited HSCs demonstrated long-term engraftment and persistence, contributing to
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