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Scientific Reports Sep 2019Human olfactory mucosa cells (hOMCs) have been transplanted to the damaged spinal cord both pre-clinically and clinically. To date mainly autologous cells have been...
Human olfactory mucosa cells (hOMCs) have been transplanted to the damaged spinal cord both pre-clinically and clinically. To date mainly autologous cells have been tested. However, inter-patient variability in cell recovery and quality, and the fact that the neuroprotective olfactory ensheathing cell (OEC) subset is difficult to isolate, means an allogeneic hOMC therapy would be an attractive "off-the-shelf" alternative. The aim of this study was to generate a candidate cell line from late-adherent hOMCs, thought to contain the OEC subset. Primary late-adherent hOMCs were transduced with a c-MycER gene that enables cell proliferation in the presence of 4-hydroxytamoxifen (4-OHT). Two c-MycER-derived polyclonal populations, PA5 and PA7, were generated and expanded. PA5 cells had a normal human karyotype (46, XY) and exhibited faster growth kinetics than PA7, and were therefore selected for further characterisation. PA5 hOMCs express glial markers (p75, S100ß, GFAP and oligodendrocyte marker O4), neuronal markers (nestin and ß-III-tubulin) and fibroblast-associated markers (CD90/Thy1 and fibronectin). Co-culture of PA5 cells with a neuronal cell line (NG108-15) and with primary dorsal root ganglion (DRG) neurons resulted in significant neurite outgrowth after 5 days. Therefore, c-MycER-derived PA5 hOMCs have potential as a regenerative therapy for neural cells.
Topics: Adult; Animals; Biomarkers; Cell Line; Coculture Techniques; Ganglia, Spinal; Genes, myc; Gentamicins; Humans; Karyotyping; Mice; Neuroblastoma; Olfactory Mucosa; Rats; Rats, Sprague-Dawley; Receptors, Estrogen; Recombinant Proteins; Sensory Receptor Cells; Tamoxifen; Transduction, Genetic; Transgenes
PubMed: 31519924
DOI: 10.1038/s41598-019-49315-6 -
Cell Communication and Signaling : CCS Aug 2019Ligands of the C-type lectin CLEC10A such as Tn and sialyl-Tn representing early intermediates of O-glycosylation are hallmarks of many human malignancies. A variety of...
BACKGROUND
Ligands of the C-type lectin CLEC10A such as Tn and sialyl-Tn representing early intermediates of O-glycosylation are hallmarks of many human malignancies. A variety of regulatory mechanisms underlying their expression are being discussed.
METHODS
CLEC10A ligands were detected in various tissues and cells using the recombinant glycan-binding domain of CLEC10A. In normal breast and endometrium, presence of ligands was correlated to the female cycle. Estrogen- and stress dependent induction of CLEC10A ligands was analyzed in MCF7 and T47D cells exposed to 4-hydroxy-tamoxifen (Tam), zeocin and hydrogen peroxide. The expression and localization of CLEC10A ligands was analyzed by Western blot and immunofluorescence. In breast cancer patients CLEC10A ligand expression and survival was correlated by Kaplan-Meyer analysis.
RESULT
We observed binding of CLEC10A in normal endometrial and breast tissues during the late phase of the female hormonal cycle suggesting a suppressive effect of female sex hormones on CLEC10A ligand expression. Accordingly, CLEC10A ligands were induced in MCF7- and T47D breast cancer cells after Tam treatment and accumulated on the cell surface and in the endosomal/lysosomal compartment. Phagocytosis experiments indicate that macrophages preferentially internalize CLEC10A ligands coated beads and Tam treated MCF7 cells. CLEC10A ligands were also expressed after the addition of zeocin and hydrogen-peroxide. Each substance induced the production of ROS indicating reactive oxygen species as a unifying mechanism of CLEC10A ligand induction. Mechanistically, increased expression of GalNAc-transferase 6 (GalNT6) and translocation of GalNT2 and GalNT6 from cis- towards trans-Golgi compartment was observed, while protein levels of COSMC and T-synthase remained unaffected. In breast cancer patients, positivity for CLEC10A staining in tumor tissues was associated with improved outcome and survival.
CONCLUSION
CLEC10A ligands are inducible by hormone depletion, 4-hydroxy-tamoxifen and agents inducing DNA damage and oxidative stress. Our results indicate that CLEC10A acts as a receptor for damaged and dead cells and may play an important role in the uptake of cell debris by macrophages and dendritic cells.
Topics: Antineoplastic Agents; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; DNA Damage; Drug Screening Assays, Antitumor; Female; HEK293 Cells; Humans; Lectins, C-Type; Ligands; MCF-7 Cells; Oxidative Stress; Polysaccharides; Recombinant Proteins; Signal Transduction; Tamoxifen
PubMed: 31455323
DOI: 10.1186/s12964-019-0420-9 -
Endocrine-related Cancer Aug 2019Breast cancer is the most prevalent malignancy and second leading cause of death in women worldwide, with hormone receptor-positive luminal breast cancers being the most...
Breast cancer is the most prevalent malignancy and second leading cause of death in women worldwide, with hormone receptor-positive luminal breast cancers being the most widespread subtype. While these tumors are generally amenable to endocrine therapy, cellular heterogeneity and acquired ability of tumor cells to undergo cell state switching makes these populations difficult to be fully targeted and eradicated through conventional methods. We have leveraged a quality-by-design (QbD) approach that integrates biological responses with predictive mathematical modeling to identify key combinations of commercially available drugs to induce estrogen receptor expression for therapeutic targeting. This technology utilizes a high level of automation through a custom-built platform to reduce bias as well as design-of-experiments methodology to minimize the experimental iterations required. Utilizing this approach, we identified a combination of clinical compounds, each at concentrations well below their efficacious dose, able to induce the expression of estrogen receptor alpha (ESR1) in hormone-positive breast cancer cells. Induction of ESR1 in luminal cells leads to chemosensitization. These findings provide proof of concept for the utility of the QbD strategy and identify a unique drug cocktail able to sensitize breast cancer cells to tamoxifen.
Topics: Antineoplastic Agents, Hormonal; Antineoplastic Combined Chemotherapy Protocols; Breast Neoplasms; Cell Line, Tumor; Cell Proliferation; Drug Resistance, Neoplasm; Estradiol; Estrogen Receptor alpha; Everolimus; Female; Humans; Hydroxamic Acids; Indazoles; MCF-7 Cells; Paclitaxel; Sulfonamides; Tamoxifen; Tumor Cells, Cultured
PubMed: 31167163
DOI: 10.1530/ERC-19-0042 -
Hematopoietic transformation in the absence of MLL1/KMT2A: distinctions in target gene reactivation.Cell Cycle (Georgetown, Tex.) Jul 2019The deregulation of hematopoietic stem cell (HSC) transcriptional networks is a common theme in acute myelogenous leukemia (AML). Chromosomal translocations that alter...
The deregulation of hematopoietic stem cell (HSC) transcriptional networks is a common theme in acute myelogenous leukemia (AML). Chromosomal translocations that alter the gene () occur in infant, childhood and adult leukemia and at the same time, wild-type MLL1 is a critical regulator of HSC homeostasis. Typically, the endogenous, wild-type (WT) MLL1 and MLL fusion oncoproteins (MLL-FPs) remain both expressed in leukemia. WT and MLL-FPs activate overlapping sets of target genes, presenting a challenge for the selective therapeutic targeting of leukemic cells. We previously demonstrated that endogenous MLL1 is not required for the maintenance of MLL-FP-driven AML but is required for normal HSC homeostasis. Here we address the role of MLL-FPs in the initiation of leukemia in the absence of endogenous MLL1. We show that loss of endogenous results in a rapid decrease in expression of shared HSC/leukemia target genes, yet MLL-AF9 restores the expression of most of these target genes in the absence of WT MLL1, with the critical exception of /. These observations underscore the sufficiency of MLL-fusion oncoproteins for initiating leukemia, but also illustrate that WT MLL1 target genes differ in their ability to be re-activated by MLL-FPs.
Topics: Animals; Antineoplastic Agents; Carcinogenesis; Female; Gene Expression Regulation, Leukemic; HEK293 Cells; Hematopoietic Stem Cell Transplantation; Hematopoietic Stem Cells; Histone-Lysine N-Methyltransferase; Humans; Leukemia, Myeloid, Acute; MDS1 and EVI1 Complex Locus Protein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myeloid-Lymphoid Leukemia Protein; Oncogene Proteins, Fusion; Tamoxifen
PubMed: 31161857
DOI: 10.1080/15384101.2019.1618642