Project description:Induced pluripotent stem cells were gene edited to introduce heterozygous (promoter deletion, start site deletion, and exon 27 frameshift) and homozygous (exon 27 frameshift) mutations. iPSCs were differentiated into cardiomyocytes for transcriptome analysis.
Project description:RBM20 heterozygous (Het) and homozygous (Homo) R636S and homozygous 8-bp deletion (which causes nonsense-mediated decay) mutations were created by genome editing in an iPS cell line generated from a healthy male patient. Those cells were differentiated into cardiomyocytes by modulating the WNT signaling pathway, and then are subjected to the lactate purification method.
Project description:RBM20 heterozygous (Het) and homozygous (Homo) R636S and homozygous 8-bp deletion (which causes nonsense-mediated decay) mutations were created by genome editing in an iPS cell line generated from a healthy male patient. Those cells were differentiated into cardiomyocytes by modulating the WNT signaling pathway, and then are subjected to the lactate purification method.
Project description:The transcription factor GATA2 plays a major role in the generation and maintenance of the hematopoietic system. In humans, heterozygous germline mutations in GATA2 often lead to a loss of function of one allele, causing GATA2 haploinsufficiency. In mice, Gata2 has an essential regulatory function in hematopoietic stem cell (HSC) generation and maintenance. However, whereas Gata2-null mice are lethal at embryonic day (E) 10.53, Gata2 heterozygous (Gata2+/-) mice survive to adulthood with normal blood values. However, mouse models thus emerged as a useful source to identify the function of GATA2 in HSC generation and fitness, they leave the mechanisms causing the different aspects of GATA2 deficiency syndrome largely undiscovered. Zebrafish have the advantage of having two GATA2 orthologues; Gata2a and Gata2b. Gata2a is expressed predominantly in the vasculature and is required for programming of the hemogenic endothelium. Gata2b is expressed in hematopoietic stem/progenitor cells (HSPCs) and homozygous deletion (gata2b-/-) redirects HSPCs differentiation bias, thus mimicking one of the GATA2 haploinsufficiency phenotypes found in patients. But patients carry heterozygous rather than homozygous GATA2 mutations, we specifically focused on how heterozygous Gata2b mutations could be mechanistically linked to erythro-myelodysplasia, a major clinical hallmark of GATA2 patients. To investigate the mechanisms of heterozygous GATA2 mutation caused GATA2 deficiency syndrome, we created a heterozygous gata2b mutation zebrafish model and sorted the entire progenitor and HSPC population including the lymphoid population from kidney marrow (KM) of WT and mutated zebrafish based on the scatter profile of flow cytometry for single-cell RNA (scRNA) sequencing.
Project description:Bulk tissue RNA-sequencing of individual 24hpf zebrafish larvae to compare the gene expression values between wild type and foxg1a nonsense mutants (heterozygous and homozygous mutants). The mutation is a 5bp deletion (32bp from canonical start codon; AAATG deleted).
Project description:The transcription factor GATA2 plays a major role in the generation and maintenance of the hematopoietic system. In humans, heterozygous germline mutations in GATA2 often lead to a loss of function of one allele, causing GATA2 haploinsufficiency. In mice, Gata2 has an essential regulatory function in hematopoietic stem cell (HSC) generation and maintenance. However, whereas Gata2-null mice are lethal at embryonic day (E) 10.53, Gata2 heterozygous (Gata2+/-) mice survive to adulthood with normal blood values. However, mouse models thus emerged as a useful source to identify the function of GATA2 in HSC generation and fitness, they leave the mechanisms causing the different aspects of GATA2 deficiency syndrome largely undiscovered. Zebrafish have the advantage of having two GATA2 orthologues; Gata2a and Gata2b. Gata2a is expressed predominantly in the vasculature and is required for programming of the hemogenic endothelium. Gata2b is expressed in hematopoietic stem/progenitor cells (HSPCs) and homozygous deletion (gata2b-/-) redirects HSPCs differentiation bias, thus mimicking one of the GATA2 haploinsufficiency phenotypes found in patients. But patients carry heterozygous rather than homozygous GATA2 mutations, we specifically focused on how heterozygous Gata2b mutations could be mechanistically linked to erythro-myelodysplasia, a major clinical hallmark of GATA2 patients. To investigate the mechanisms of heterozygous GATA2 mutation caused GATA2 deficiency syndrome, we created a heterozygous gata2b mutation zebrafish model and sorted the entire progenitor and HSPC population including the lymphoid population from kidney marrow (KM) of WT and mutated zebrafish based on the scatter profile of flow cytometry for single-nucleus ATAC (snATAC) sequencing.
Project description:To study Hnrnpu function during cortical development we preformed single-cell RNA sequencing (scRNA-seq) on mouse neutrosphere cultures derived from E13 ICR cortices, 24 hours following treatment with CRISPR/CAS9 and two sgRNA sequences targeting mouse Hnrnpu. In parallel, we analyzed bulk RNA-seq from dissected cortices of E13 embryos in which Hnrnpu conditional truncation in the telencephalon was driven by Emx1::Cre. Additionally, we analyzed MARS-Seq trancriptomal profiles of cortices of E13 embryos carrying homozygous heterozygous or wildtype alleles of Hnrnpu truncation (Emx1::Cre driver) in combination with homozygous or heterozygous conditional deletion of Tp53
Project description:Interventions: Administraion of CPT-11 is twice for 4 weeks on days 1 and 15. CPT-11 is reconstituted in >=250 mL of normal saline or 5% dextrose in water and infuse 90min on day 1 for pharmacokinetics, and 90min over on day15. CPT-11 adjusted dosage is determined from 50,75,100,125 or 150mg/sqm in the heterozygous group and the homozygous group by continual reassessment method. CPT-11 dosage is fixed at 150mg/sqm in the wild group.Definision of UGT1A1 polymorphisms groups: The homozygous group is patient with homozygous genotype of UGT1A1*28/*28 or UGT1A1*6/*6, with combined heterozygous genotypes of UGT1A1*28 and UGT1A1*6. The heterozygous group is patient with heterozygous genotype of either UGT1A1*28 or UGT1A1*6. The wild group is patients with no UGT1A1*28 and UGT1A1*6 mutation.
Primary outcome(s): Maximum tolerated dose for the heterozygous group and the homozygous group, respectively. Incidence rate of dose limiting toxicities for the wild group.
Study Design: Single arm Non-randomized
Project description:Interventions: Patients receive FOLFIRI with bevacizumab fixed 5mg/kg. (Treatment will be continued unless the disease progression, unacceptable toxicity, or consent withdrawal.) Definision of UGT1A1 polymorphisms groups: The homozygous group is patient with homozygous genotype of UGT1A1*28/*28 or UGT1A1*6/*6, with combined heterozygous genotypes of UGT1A1*28 and UGT1A1*6. The heterozygous group is patient with heterozygous genotype of either UGT1A1*28 or UGT1A1*6. The wild group is patients with no UGT1A1*28 and UGT1A1*6 mutation. CPT-11 dosage is wild and heterozygous:CPT-11 150mg/m2 homozygous:CPT-11 100mg/m2
Primary outcome(s): 1)incidence of adverse events 2)frequency of severe toxicity
Study Design: Single arm Non-randomized
