ABSTRACT
Cellular heterogeneity, for example, the intratumoral coexistence of cancer cells with and without stem cell characteristics, represents a potential root of therapeutic resistance and a significant challenge for modern drug development in glioblastoma (GBM). We propose here that activation of the innate immune system by stimulation of innate immune receptors involved in antiviral and antitumor responses can similarly target different malignant populations of glioma cells. We used short-term expanded patient-specific primary human GBM cells to study the stimulation of the cytosolic nucleic acid receptors melanoma differentiation-associated gene 5 (MDA5) and retinoic acid-inducible gene I (RIG-I). Specifically, we analyzed cells from the tumor core versus "residual GBM cells" derived from the tumor resection margin as well as stem cell-enriched primary cultures versus specimens without stem cell properties. A portfolio of human, nontumor neural cells was used as a control for these studies. The expression of RIG-I and MDA5 could be induced in all of these cells. Receptor stimulation with their respective ligands, p(I:C) and 3pRNA, led to in vitro evidence for an effective activation of the innate immune system. Most intriguingly, all investigated cancer cell populations additionally responded with a pronounced induction of apoptotic signaling cascades revealing a second, direct mechanism of antitumor activity. By contrast, p(I:C) and 3pRNA induced only little toxicity in human nonmalignant neural cells. Granted that the challenge of effective central nervous system (CNS) delivery can be overcome, targeting of RIG-I and MDA5 could thus become a quintessential strategy to encounter heterogeneous cancers in the sophisticated environments of the brain.
Subject(s)
Antineoplastic Agents/pharmacology , Cytosol/immunology , DEAD-box RNA Helicases/immunology , Glioblastoma/drug therapy , Glioblastoma/immunology , Apoptosis/drug effects , Apoptosis/genetics , Apoptosis/immunology , Brain Neoplasms/drug therapy , Brain Neoplasms/genetics , Brain Neoplasms/immunology , Brain Neoplasms/metabolism , Cell Line, Tumor , Cytosol/drug effects , Cytosol/metabolism , DEAD Box Protein 58 , DEAD-box RNA Helicases/genetics , DEAD-box RNA Helicases/metabolism , Glioblastoma/genetics , Glioblastoma/metabolism , Humans , Immunity, Innate/drug effects , Immunity, Innate/genetics , Immunity, Innate/immunology , Interferon-Induced Helicase, IFIH1 , Ligands , Receptors, Immunologic , Signal Transduction/drug effects , Stem Cells/drug effects , Stem Cells/immunology , Stem Cells/metabolismABSTRACT
Hyaluronan and its receptor CD44 are known to contribute to the invasive growth of different tumors of the central nervous system. It is not known, however, if CD44 is sufficient to activate invasive growth into the brain tissue. This study examines how CD44 regulates the motility and invasive growth of B35 neuroblastoma cells into a hyaluronan-rich environment. A comprehensive experimental approach was used encompassing biochemical techniques, single molecule microscopy, correlative confocal and scanning electron microscopy, morphometry of cellular extensions, live-cell imaging and tracking, transplantation onto organotypic brain slices, two-photon imaging and invasion assays. We found that CD44-GFP fusion protein was localized in filopodia and in focal bleb-like protrusions where it provided binding sites for hyaluronan. Transient expression of CD44-GFP was sufficient to increase the length of filopodia, to enhance cell migration and to promote invasive growth into hyaluronan-rich brain tissue. Thus, CD44 controls molecular devices localized in filopodia and bleb-like specializations of the cell surface that enhance cell migration and invasive growth.
