Biology subjects

Shoichet, M. S.

Publications and source records attributed to Shoichet, M. S..

4 recordsLinked to original sources

bioRxiv2025

Identification of Injury-Response State Determinants in Glioblastoma Stem Cells

Glioblastoma (GBM) is a common and highly lethal type of primary brain tumor in adults. Therapeutic failure is partly attributed to a fraction of Glioblastoma Stem Cells (GSCs) that show high levels of heterogeneity and plasticity. GSCs exist in a transcriptional gradient between two states: Developmental (Dev) and Injury Response (IR) in which IR-GSCs exhibit more invasive behaviors. While previous studies have identified fitness genes in GSCs, the genes required to establish and maintain the Dev and IR states remain poorly defined. To identify the regulators of the IR GSC state, we performed a phenotypic genome-wide CRISPR-Cas9 knockout (KO) screen in patient-derived GSCs based on cell surface expression of the IR marker CD44. Notably, we found that perturbations of the histone acetyltransferase EP300 in IR GSCs led to decreased CD44 cell surface expression, significant downregulation of gene expression signatures associated with the IR state, and to decreased self-renewal and invasion. Furthermore, genetic targeting of Ep300 in a mouse GBM model delayed tumor initiation and/or progression. Collectively, our results establish EP300 as a regulator of the IR state in GSCs and provide a mechanistic basis for its therapeutic targeting in GBM. SignificanceA genome-wide phenotypic CRISPR-Cas9 screen in a patient-derived Glioblastoma Stem Cell line identified the genes required to maintain the Injury-Response cellular state, with a focus on the histone acetyl transferase gene EP300. This study suggests how therapeutic targeting of cellular state could reduce the aggressiveness of GBM tumors.

cancer biology↗

bioRxiv2022

Viscoelastic Notch Signaling Hydrogel Induces Liver Bile Duct Organoid Growth and Morphogenesis

Cholangiocyte organoids can be used to model liver biliary disease; however, both a defined matrix in which to emulate cholangiocyte self-assembly and the mechano-transduction pathways involved therein remain elusive. We designed a series of defined viscoelastic hyaluronan hydrogels in which to culture primary cholangiocytes and found that by mimicking the stress relaxation rate of liver tissue, we could induce cholangiocyte organoid growth and significantly increase expression of Yes-associated protein (YAP) target genes. Strikingly, inhibition of matrix metalloproteinases (MMPs) did not significantly affect organoid growth in 3D culture, suggesting that mechanical remodeling of the viscoelastic microenvironment - and not MMP-mediated degradation - is key to cholangiocyte organoid growth. By immobilizing jagged1 to the hyaluronan, stress relaxing hydrogel, self-assembled bile duct structures formed in organoid culture, indicating the synergistic effects of Notch signaling and viscoelasticity. By uncovering critical roles of hydrogel viscoelasticity, YAP signaling and Notch activation, we controlled cholangiocyte organogenesis, thereby paving the way for their use in disease modeling and/or transplantation.

bioengineering↗

bioRxiv2021

Three-dimensional drug screen identifies HDAC inhibitors as therapeutic agents in mTORC1-driven lymphangioleiomyomatosis

Lymphangioleiomyomatosis (LAM) is a rare disease involving cystic lung destruction by invasive LAM cells. These cells harbor loss-of-function mutations in TSC2, conferring constitutive mTORC1 signaling. Rapamycin is the only clinically approved disease-modifying treatment, but its action is cytostatic and disease progresses upon its withdrawal. There is a critical need to identify novel agents that prevent the invasive phenotype and/or eradicate the neoplastic LAM cells. Here, we employed novel cellular and extracellular models to screen for candidate therapeutics in a physiologically relevant setting. We observed that lung-mimetic hydrogel culture of pluripotent stem cell-derived diseased cells more faithfully recapitulates human LAM biology compared to conventional culture on two-dimensional tissue culture plastic. Leveraging our culture system, we conducted a three-dimensional drug screen using a custom 800-compound library, tracking cytotoxicity and invasion modulation phenotypes at the single cell level. We identified histone deacetylase (HDAC) inhibitors as a group of anti-invasive agents that are also selectively cytotoxic towards TSC2-/- cells. Unexpectedly, we observed that next generation ATP-competitive mTORC1/2 inhibitors potentiate invasion. We determined anti-invasive effects of HDAC inhibitors to be independent of genotype, while selective cell death is mTORC1-dependent and mediated by apoptosis. Drug performance was subsequently evaluated at the single cell level in zebrafish xenografts. We observed consistent therapeutic efficacy in vivo at equivalent concentrations to those used in vitro, substantiating HDAC inhibitors as potential therapeutic candidates for pursuit in patients with LAM. One Sentence SummaryWe performed a drug screen in 3D and discovered HDAC inhibitors exhibit therapeutic efficacy in models of the lung disease lymphangioleiomyomatosis.

cell biology↗

bioRxiv2018

Muscle stem cell intramuscular delivery within hyaluronan methylcellulose improves engraftment efficiency and dispersion

Adult skeletal muscle tissue harbors the capacity for self-repair due to the presence of tissue resident muscle stem cells (MuSCs). Advances in the area of prospective MuSC isolation demonstrated the potential of cell transplantation therapy as a regenerative medicine strategy to restore strength and long-term regenerative capacity to aged, injured, or diseased skeletal muscle tissue. However, cell loss during ejection, limits to post-injection proliferation, and poor donor cell dispersion distal to the injection site are amongst hurdles to overcome to maximize MuSC transplant impact. Here, we assess a physical blend of hyaluronan and methylcellulose (HAMC) as a bioactive, shear thinning hydrogel cell delivery system to improve MuSC transplantation efficiency. Using in vivo transplantation studies, we found that the HAMC delivery system results in a >45% increase in the number of donor-derived fibers as compared to saline delivery. Furthermore, we observed a significant improvement in donor fiber dispersion when transplanted MuSCs were delivered in the HAMC hydrogel. Studies to assess primary myoblast and MuSC viability in HAMC culture revealed no differences compared to the media control even when the cells were first ejected through a syringe and needle or exposed to regenerating skeletal muscle extract to mimic the transplantation procedure. However, when we quantified absolute numbers, we found that more cells pass through the syringe and needle when delivered in HAMC. Culture in HAMC also increased the proportion of MuSCs in cell cycle, via a CD44-independent mechanism. An effect on myoblast proliferation was not observed, suggesting a hierarchical effect. Finally, a series of transplant studies indicated that HAMC delivery does not influence passive cell clearance or alter the host immune response, but instead may serve to support in vivo expansion by delaying differentiation following transplant. Therefore, we conclude that MuSC engraftment efficacy is improved by delivering the therapeutic cell population within HAMC.

bioengineering↗