Development of model systems for b-thalassaemia

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Interplay between α‐thalassemia and β‐hemoglobinopathies: Translating genotype–phenotype relationships into therapies

Abstract α‐Thalassemia represents one of the most important genetic modulators of β‐hemoglobinopathies. During this last decade, the ongoing interest in characterizing genotype–phenotype relationships has yielded incredible insights into α‐globin gene regulation and its impact on β‐hemoglobinopathies. In this review, we provide a holistic update on α‐globin gene expression stemming from DNA to RNA to protein, as well as epigenetic mechanisms that can impact gene expression and potentially influence phenotypic outcomes. Here, we highlight defined α‐globin targeted strategies and rationalize the use of distinct molecular targets based on the restoration of balanced α/β‐like globin chain synthesis. Considering the therapies that either increase β‐globin synthesis or reactivate γ‐globin gene expression, the modulation of α‐globin chains as a disease modifier for β‐hemoglobinopathies still remains largely uncharted in clinical studies

A Cas9 Variant for Efficient Generation of Indel-Free Knockin or Gene-Corrected Human Pluripotent Stem Cells

While Cas9 nucleases permit rapid and efficient generation of gene-edited cell lines, the CRISPR-Cas9 system can introduce undesirable “on-target” mutations within the second allele of successfully modified cells via non-homologous end joining (NHEJ). To address this, we fused the Streptococcus pyogenes Cas9 (SpCas9) nuclease to a peptide derived from the human Geminin protein (SpCas9-Gem) to facilitate its degradation during the G1 phase of the cell cycle, when DNA repair by NHEJ predominates. We also use mRNA transfection to facilitate low and transient expression of modified and unmodified versions of Cas9. Although the frequency of homologous recombination was similar for SpCas9-Gem and SpCas9, we observed a marked reduction in the capacity for SpCas9-Gem to induce NHEJ-mediated indels at the target locus. Moreover, in contrast to native SpCas9, we demonstrate that transient SpCas9-Gem expression enables reliable generation of both knockin reporter cell lines and genetically repaired patient-specific induced pluripotent stem cell lines free of unwanted mutations at the targeted locus

Intranasal immunization with yeast-expressed 19 kD carboxyl-terminal fragment of Plasmodium yoelii merozoite surface protein-1 (yMSP119) induces protective immunity to blood stage malaria infection in mice

Variable protection against malaria blood-stage infection has been demonstrated in mice following parenteral immunization with the highly conserved 19 kD carboxylterminal fragment of the merozoite surface protein-1 (MSP) using CFA/IFA and other adjuvants. Here we show that intranasal immunization of BALB/C mice with yeast expressed Plasmodium yoelii MSP1 plus a mixture of native and recombinant cholera toxin B subunit, could induce serum MSP1-specific antibodies at titres ranging from 20,000 to 2.56,000. The Ig subclass responses were predominantly G1 and G2b. Intranasal immunization led to protection following challenge (peak parasitaemi