Project description:In order to determine the transcriptional effect of Ikzf2 overexpression in mammalian auditory hair cells, mouse cochleae were transfected with either a control GFP or a Ikzf2 virus between postnatal day 1 and 3 (P1-3) and then harvested for single cell gene expression profiling at postnatal day 8 (P8) on the 10X Genomics Single Cell 3' v2 platform.
Project description:Mammalian cochlear outer hair cells (OHCs) are essential for hearing. Severe hearing impairment follows OHC degeneration. Previous attempts at regenerating new OHCs from cochlear supporting cells (SCs) have been unsuccessful, notably lacking expression of the key OHC motor protein, Prestin. Thus, regeneration of Prestin+ OHCs represented a barrier to restoring auditory function in vivo. Here, we reported the successful in vivo conversion of adult mouse cochlear SCs into Prestin+ OHC-like cells through the concurrent induction of two key transcriptional factors known to be necessary for OHC development: Atoh1 and Ikzf2. Single cell RNA sequencing reveals the upregulation of 729 OHC genes and downregulation of 331 SC genes in OHC-like cells. The resulting differentiation status of these OHC-like cells was much more advanced than previously achieved. This study thus established an efficient approach to induce the regeneration of Prestin+ OHCs, paving the way for in vivo cochlear repair via SC transdifferentiation.
Project description:IKZF2 is an important nuclear matrix protein and plays a pivotal role in T cell development and differentiation, while its expression and function in Cutaneous T cell lymphoma (CTCL) remain ambiguous. Our study aimed to investigate the expression pattern, biological function of IKZF2 in a large clinical cohort. IKZF2 is specifically over-expressed in malignant T cells with MF tumor-stage. In order to explore the function of IKZF2 in the pathogenesis of CTCL, transcriptome sequencing was performed among Hut78 cells transfected with shRNAs targeting IKZF2 (shIKZF2) and scrambled shRNA(sh0) to investigate genes regulated by IKZF2 in CTCL cells .
Project description:Acute myeloid leukemia (AML) is a hematologic malignancy for which several epigenetic regulators have been identified as therapeutic targets. Here we report the development of cereblon-dependent degraders of IKZF2 and casein kinase 1 alpha (CK1α) termed DEG-35 and DEG-77. We utilized a structure-guided approach to develop DEG-35 as a nanomolar degrader of IKZF2, a hematopoietic specific transcription factor that contributes to myeloid leukemogenesis. DEG-35 possesses additional substrate specificity for the therapeutically relevant target CK1α which was identified through unbiased proteomics and a PRISM screen assay. Degradation of IKZF2 and CK1α blocks cell growth and induces myeloid differentiation in AML cell lines through CK1α–p53- and IKZF2-dependent pathways. Target degradation by DEG-35 or the analog DEG-77 delays leukemia progression in murine and human AML mice models. Overall, we provide a strategy for multi-targeted degradation of IKZF2/CK1α to enhance efficacy against AML that may be expanded to additional targets and indications.
Project description:Lenalidomide achieves its therapeutic efficacy by recruiting and removing proteins of therapeutic interest through the E3 ligase substrate adapter cereblon. Here, we report the rational design and characterization of 81 cereblon ligands for their ability to degrade the transcription factor Helios (IKZF2) and casein kinase 1 alpha (CK1α) in acute myeloid leukemia MOLM-13 cells. Using a structure-based approach, we identified a key naphthamide scaffold that depleted both intended targets. Structure-activity relationship studies for degradation of the desired targets over other targets (IKZF1, GSPT1) afforded an initial lead compound, termed DEG-35. A subsequent scaffold replacement campaign informed by degradation profiles against a panel of substrates identified DEG-77, which selectively degrades IKZF2 and CK1α, and possesses suitable pharmacokinetic properties, solubility, and selectivity for in vivo studies. Finally, we show that DEG-77 has antiproliferative activity in diffuse large B cell lymphoma (DLBCL) cell line OCI-LY3 and ovarian cancer cell line A2780, indicating that these dual degraders and their targets may have efficacy against additional cancer types.
Project description:Deprivation of peripheral nerve input by cochlear removal in young mice results in dramatic neuron death in the cochlear nucleus (CN). The same manipulation in older mice does not result in significant loss. The molecular basis of this critical period of vulnerability remains largely unknown. Here we identified genes regulated at early time points after cochlear removal at ages when neurons are vulnerable (postnatal day (P)7) or invulnerable (P21) to this challenge. Afferent deprivation regulated very different sets of genes at P7 and P21. These genes showed a variety of functions at both ages, but surprisingly there was no net increase in pro-apoptotic genes at P7. A large set of upregulated immune-related genes was identified at P21. Experiment Overall Design: Mice received unilateral cochlear removals. At 6, 12, 24, and 48 hours after surgery, the CN ipsilateral and contralateral were removed, and RNA isolated from separate pools of tissue for each replicate. Deafferented CN were compared to age-matched and time-matched contralateral, control CN to identify genes regulated by cochlear removal at age P7 and P21.