Project description:C-to-T base editing mediated by CRISPR/Cas9 base editors (BEs) needs a G/C-rich PAM and the editing fidelity is compromised by unwanted indels and non-C-to-T substitutions. We developed CRISPR/Cpf1-based BEs to recognize a T-rich PAM and induce efficient C-to-T editing with few indels and/or non-C-to-T substitutions. The requirement of editing fidelity in therapeutic-related trials necessitates the development of CRISPR/Cpf1-based BEs, which also facilitates base editing in A/T-rich regions.
Project description:The targeting range of CRISPR-Cas9 base editors (BEs) is limited by their G/C-rich PAM sequences. To overcome this limitation, we developed a CRISPR/Cpf1-based BE by fusing the rat cytosine deaminase APOBEC1 to a catalytically inactive version of Lachnospiraceae bacterium Cpf1. The base editor recognizes a T-rich PAM sequence and converts C to T in human cells with low levels of indels, non-C-to-T substitutions and off-target editing.
Project description:Interventions: Blood samples(10 points) are collected after the first administration of capecitabine for pharmacokinetic analysis and cytidine deaminase activity measurement.
Primary outcome(s): To evaluate the correlation AUC of 5-DFUR/AUC of 5-DFCR ratio and cytidine deaminase activity.
Study Design: Single arm Non-randomized
Project description:C-to-T base editing mediated by CRISPR/Cas9 base editors (BEs) needs a GC-rich PAM and the editing fidelity is compromised by unwanted indels and non-C-to-T substitutions. We developed CRISPR/Cpf1-based BEs to recognize a T-rich PAM and induce efficient C-to-T editing with few indels and/or non-C-to-T substitutions. The requirement of editing fidelity in therapeutic-related trials necessitates the development of CRISPR/Cpf1-based BEs which also facilitates base editing in AT-rich regions.
Project description:We report the analysis of genome wide distribution of activation induced cytidine deaminase by ChIP-seq in Human B cells. The resultant distribution of AID has then been compared to various histone modifications and transcription factors to shed insights into the mechanisms that target this enzyme.
Project description:This study evaluates the similarity of adaptive immune mechanisms between jawless and jawed vertebrates using lamprey cytidine deaminase (CDA). We identified the ancestral gene Lr-CDAs of the AID/APOBEC deaminase family and evaluated its biological function in vivo. Lr-CDA1 deletion affected the assembly of three types of variable lymphocyte receptors (VLRs). We identified a switch-like region in lamprey gVLRs bound to Lr-CDAs, which upon repression, downregulated VLRB expression. Overall, we propose that lampreys have an early form of class switch recombination (CSR) that is mediated by Lr-CDAs and acts on gVLRs, affecting the assembly, maturation, and diversity regulation of VLR genes in Lethenteron reissneri. This CSR process in lampreys is linked to tumorigenesis and chromosomal translocation markers via Lr-CDAs.