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1.
Sci Adv ; 10(9): eadj5107, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38427725

ABSTRACT

Cell fate decisions are achieved with gene expression changes driven by lineage-specific transcription factors (TFs). These TFs depend on chromatin remodelers including the Brahma-related gene 1 (BRG1)-associated factor (BAF) complex to activate target genes. BAF complex subunits are essential for development and frequently mutated in cancer. Thus, interrogating how BAF complexes contribute to cell fate decisions is critical for human health. We examined the requirement for the catalytic BAF subunit BRG1 in neural progenitor cell (NPC) specification from human embryonic stem cells. During the earliest stages of differentiation, BRG1 was required to establish chromatin accessibility at neuroectoderm-specific enhancers. Depletion of BRG1 dorsalized NPCs and promoted precocious neural crest specification and enhanced neuronal differentiation. These findings demonstrate that BRG1 mediates NPC specification by ensuring proper expression of lineage-specific TFs and appropriate activation of their transcriptional programs.


Subject(s)
Chromatin , Neural Plate , Humans , Chromatin/genetics , DNA Helicases/genetics , DNA Helicases/metabolism , Neural Plate/metabolism , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Transcription Factors/metabolism
2.
Cell Rep ; 36(4): 109429, 2021 07 27.
Article in English | MEDLINE | ID: mdl-34320344

ABSTRACT

Patient-derived tumor organoids (TOs) are emerging as high-fidelity models to study cancer biology and develop novel precision medicine therapeutics. However, utilizing TOs for systems-biology-based approaches has been limited by a lack of scalable and reproducible methods to develop and profile these models. We describe a robust pan-cancer TO platform with chemically defined media optimized on cultures acquired from over 1,000 patients. Crucially, we demonstrate tumor genetic and transcriptomic concordance utilizing this approach and further optimize defined minimal media for organoid initiation and propagation. Additionally, we demonstrate a neural-network-based high-throughput approach for label-free, light-microscopy-based drug assays capable of predicting patient-specific heterogeneity in drug responses with applicability across solid cancers. The pan-cancer platform, molecular data, and neural-network-based drug assay serve as resources to accelerate the broad implementation of organoid models in precision medicine research and personalized therapeutic profiling programs.


Subject(s)
Neoplasms/pathology , Organoids/pathology , Precision Medicine , Cell Proliferation , Drug Screening Assays, Antitumor , Female , Fluorescence , Genomics , HLA Antigens/genetics , Humans , Loss of Heterozygosity , Male , Middle Aged , Models, Biological , Neoplasms/genetics , Neural Networks, Computer , Transcriptome/genetics
3.
Elife ; 82019 04 29.
Article in English | MEDLINE | ID: mdl-31033435

ABSTRACT

The SWI/SNF complex is a critical regulator of pluripotency in human embryonic stem cells (hESCs), and individual subunits have varied and specific roles during development and in diseases. The core subunit SMARCB1 is required for early embryonic survival, and mutations can give rise to atypical teratoid/rhabdoid tumors (AT/RTs) in the pediatric central nervous system. We report that in contrast to other studied systems, SMARCB1 represses bivalent genes in hESCs and antagonizes chromatin accessibility at super-enhancers. Moreover, and consistent with its established role as a CNS tumor suppressor, we find that SMARCB1 is essential for neural induction but dispensable for mesodermal or endodermal differentiation. Mechanistically, we demonstrate that SMARCB1 is essential for hESC super-enhancer silencing in neural differentiation conditions. This genomic assessment of hESC chromatin regulation by SMARCB1 reveals a novel positive regulatory function at super-enhancers and a unique lineage-specific role in regulating hESC differentiation.


Subject(s)
Embryonic Stem Cells/metabolism , Enhancer Elements, Genetic/genetics , SMARCB1 Protein/genetics , SMARCB1 Protein/metabolism , Cell Differentiation/genetics , Child , Chromatin/genetics , Endoderm , Gene Knockdown Techniques , Genes, Tumor Suppressor , Humans , Mesoderm , Mutation/genetics , Rhabdoid Tumor/genetics
5.
Stem Cells ; 33(10): 2925-35, 2015 Oct.
Article in English | MEDLINE | ID: mdl-26119756

ABSTRACT

MicroRNAs and chromatin remodeling complexes represent powerful epigenetic mechanisms that regulate the pluripotent state. miR-302 is a strong inducer of pluripotency, which is characterized by a distinct chromatin architecture. This suggests that miR-302 regulates global chromatin structure; however, a direct relationship between miR-302 and chromatin remodelers has not been established. Here, we provide data to show that miR-302 regulates Brg1 chromatin remodeling complex composition in human embryonic stem cells (hESCs) through direct repression of the BAF53a and BAF170 subunits. With the subsequent overexpression of BAF170 in hESCs, we show that miR-302's inhibition of BAF170 protein levels can affect the expression of genes involved in cell proliferation. Furthermore, miR-302-mediated repression of BAF170 regulates pluripotency by positively influencing mesendodermal differentiation. Overexpression of BAF170 in hESCs led to biased differentiation toward the ectoderm lineage during EB formation and severely hindered directed definitive endoderm differentiation. Taken together, these data uncover a direct regulatory relationship between miR-302 and the Brg1 chromatin remodeling complex that controls gene expression and cell fate decisions in hESCs and suggests that similar mechanisms are at play during early human development.


Subject(s)
Cell Differentiation/genetics , Chromatin Assembly and Disassembly/genetics , DNA Helicases/genetics , Embryonic Stem Cells/metabolism , MicroRNAs/genetics , Nuclear Proteins/genetics , Transcription Factors/genetics , Actins/genetics , Actins/metabolism , Cell Proliferation/genetics , Chromatin/genetics , Chromosomal Proteins, Non-Histone/genetics , Chromosomal Proteins, Non-Histone/metabolism , DNA Helicases/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Endoderm/growth & development , Humans , MicroRNAs/metabolism , Nuclear Proteins/metabolism , Pluripotent Stem Cells , Transcription Factors/metabolism
6.
Cleft Palate Craniofac J ; 51(1): 110-4, 2014 Jan.
Article in English | MEDLINE | ID: mdl-23638914

ABSTRACT

OBJECTIVE: While SEX-determining region Y-Box 2 (SOX2) mutations are typically recognized as yielding ocular and central nervous system abnormalities, they have also been associated with other craniofacial defects. To elucidate the genesis of the latter, Sox2 hypomorphic (Sox2(HYP)) mice were examined, with particular attention to secondary palatal development. RESULTS: Clefts of the secondary palate were found to be highly penetrant in Sox2(HYP) mice. The palatal clefting occurred in the absence of mandibular hypoplasia and resulted from delayed or failed shelf elevation. CONCLUSIONS: Sox2 hypomorphism can result in clefting of the secondary palate, an effect that appears to be independent of mandibular hypoplasia and is thus expected to result from an abnormality that is inherent to the palatal shelves and/or their progenitor tissues. Further clinical attention relative to SOX2 mutations as a basis for secondary palatal clefts appears warranted.


Subject(s)
Cleft Palate/genetics , SOXB1 Transcription Factors/deficiency , SOXB1 Transcription Factors/genetics , Alleles , Animals , Disease Models, Animal , Genotype , Mice
7.
Development ; 140(7): 1445-56, 2013 Apr.
Article in English | MEDLINE | ID: mdl-23462474

ABSTRACT

Within discrete regions of the developing mammalian central nervous system, small subsets of glia become specialized to function as neural stem cells. As a result of their self-renewal and neurogenic capacity, these cells later serve to replenish neurons and glia during persistent or injury-induced adult neurogenesis. SOX2, an HMG box transcription factor, plays an essential role in the maintenance of both embryonic and adult neural progenitors. It is unclear, however, which biological mechanisms regulated by SOX2 are required for neural stem cell maintenance. In this study, we address this question through genetic analysis of SOX2 function in differentiating postnatal Müller glia, a cell type that maintains neurogenic capacity in the adult retina. By utilizing molecular analysis and real-time imaging, we show that two progenitor characteristics of nascent Müller glia - their radial morphology and cell cycle quiescence - are disrupted following conditional genetic ablation of Sox2 in the mouse postnatal retina, leading to Müller cell depletion and retinal degeneration. Moreover, we demonstrate that genetic induction of the Notch signaling pathway restores Müller glial cell identity to Sox2 mutant cells, but does not secure their quiescent state. Collectively, these results uncouple the roles of SOX2 and the Notch signaling pathway in the postnatal retina, and uncover a novel role for SOX2 in preventing the depletion of postnatal Müller glia through terminal cell division.


Subject(s)
Neuroglia/physiology , Retina/cytology , SOXB1 Transcription Factors/physiology , Stem Cells/physiology , Animals , Animals, Newborn , Cell Cycle Checkpoints/genetics , Cell Differentiation/genetics , Cell Differentiation/physiology , Cell Proliferation , Cells, Cultured , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neurogenesis/genetics , Neurogenesis/physiology , Neuroglia/cytology , Neuroglia/metabolism , Retina/metabolism , SOXB1 Transcription Factors/genetics , SOXB1 Transcription Factors/metabolism , Stem Cells/metabolism
8.
Mech Dev ; 129(1-4): 1-12, 2012.
Article in English | MEDLINE | ID: mdl-22522080

ABSTRACT

Haploinsufficiency for the HMG-box transcription factor SOX2 results in abnormalities of the human ventral forebrain and its derivative structures. These defects include anophthalmia (absence of eye), microphthalmia (small eye) and hypothalamic hamartoma (HH), an overgrowth of the ventral hypothalamus. To determine how Sox2 deficiency affects the morphogenesis of the ventral diencephalon and eye, we generated a Sox2 allelic series (Sox2(IR), Sox2(LP), and Sox2(EGFP)), allowing for the generation of mice that express germline hypomorphic levels (<40%) of SOX2 protein and that faithfully recapitulate SOX2 haploinsufficient human phenotypes. We find that Sox2 hypomorphism significantly disrupts the development of the posterior hypothalamus, resulting in an ectopic protuberance of the prechordal floor, an upregulation of Shh signaling, and abnormal hypothalamic patterning. In the anterior diencephalon, both the optic stalks and optic cups (OC) of Sox2 hypomorphic (Sox2(HYP)) embryos are malformed. Furthermore, Sox2(HYP) eyes exhibit a loss of neural potential and coloboma, a common phenotype in SOX2 haploinsufficient humans that has not been described in a mouse model of SOX2 deficiency. These results establish for the first time that germline Sox2 hypomorphism disrupts the morphogenesis and patterning of the hypothalamus, optic stalk, and the early OC, establishing a model of the development of the abnormalities that are observed in SOX2 haploinsufficient humans.


Subject(s)
Eye Abnormalities/genetics , Hypothalamus/abnormalities , SOXB1 Transcription Factors/genetics , Animals , Diencephalon/abnormalities , Diencephalon/pathology , Disease Models, Animal , Embryo, Mammalian/abnormalities , Embryo, Mammalian/pathology , Haploinsufficiency , Humans , Hypothalamus/pathology , Mice , Mice, Transgenic , Microscopy, Electron, Scanning , Organ Specificity , SOXB1 Transcription Factors/deficiency
9.
Surg Oncol Clin N Am ; 16(4): 873-900, x, 2007 Oct.
Article in English | MEDLINE | ID: mdl-18022550

ABSTRACT

Because chemotherapy is standard in the treatment of colorectal cancer, it is important to demonstrate whether immunizations may be given to patients receiving systemic chemotherapy. Although some studies have demonstrated immune responses in patients with metastatic colorectal carcinoma who failed standard chemotherapy, the setting of minimal residual disease may be the preferred setting for cancer vaccines. It may be important to choose antigens that have functions important to the cancer cell. The best adjuvant is not well established and may depend on the type of immune response desired. The immune system is "programmed" to down-regulate immune responses once they have become activated to avoid the development of autoimmune disease.


Subject(s)
Cancer Vaccines/therapeutic use , Colorectal Neoplasms/therapy , Immunotherapy , Cancer Vaccines/immunology , Colonic Neoplasms/drug therapy , Colonic Neoplasms/immunology , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/immunology , Humans , Immunotherapy/methods , Immunotherapy/trends , Rectal Neoplasms/drug therapy , Rectal Neoplasms/immunology , Vaccination
10.
Expert Opin Biol Ther ; 7(8): 1245-56, 2007 Aug.
Article in English | MEDLINE | ID: mdl-17696822

ABSTRACT

Breaking immune tolerance against tumor self-antigens is presently an area of intense research in the design of cancer therapies. One possible method to enhance immune system activation against tumor antigens is by blocking the inhibitory co-stimulatory signals mediated by cytotoxic T lymphocyte antigen 4, (CTLA-4) expressed on activated T cells. The fully human monoclonal antibodies that are directed against human CTLA-4, ipilimumab (Medarex/Bristol-Myers Squibb) and CP-675,206 (Pfizer/Abgenix, now Amgen), have demonstrated activity against metastatic melanoma, hormone refractory prostate cancer and other malignancies. They have also uncovered unusual immune-related adverse events manifesting as self-limiting inflammatory reactions of the bowel, skin and pituitary. This article reviews preclinical development and data generated from Phase I, II and III studies with regard to the end points reported and immune-related adverse events.


Subject(s)
Antibodies, Blocking/therapeutic use , Antigens, CD/immunology , Antigens, CD/therapeutic use , Antigens, Differentiation/immunology , Antigens, Differentiation/therapeutic use , Randomized Controlled Trials as Topic , Animals , CTLA-4 Antigen , Humans , Randomized Controlled Trials as Topic/methods
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