Development of 3D culture models of plexiform neurofibroma and initial application for phenotypic characterization and drug screening

• Published in: Experimental neurology 2018-01, Vol.299 (Pt B), p.289-298
• Main Authors: Kraniak, Janice M., Chalasani, Anita, Wallace, Margaret R., Mattingly, Raymond R.
• Format: Article
• Language: English
• Subjects: 3D culture model; Antineoplastic Agents - pharmacology; Benzimidazoles - pharmacology; Bone Morphogenetic Protein 2 - antagonists & inhibitors; Cell Culture Techniques; Cells, Cultured; Dose-Response Relationship, Drug; Drug Resistance, Neoplasm; Drug Screening Assays, Antitumor - methods; Genes, Neurofibromatosis 1; Genes, Reporter; Humans; Luminescent Proteins - analysis; Luminescent Proteins - genetics; MAP Kinase Kinase 1 - antagonists & inhibitors; Matrix proteolysis; Molecular Targeted Therapy; Neurofibroma, Plexiform - drug therapy; Neurofibroma, Plexiform - genetics; Neurofibroma, Plexiform - pathology; Neurofibromatosis 1 - pathology; NF1; Phenotype; Plexiform neurofibroma; Podophyllotoxin - analogs & derivatives; Podophyllotoxin - pharmacology; Protein Kinase Inhibitors - pharmacology; Pyrazoles - pharmacology; Pyrimidines - pharmacology; Receptor, IGF Type 1 - antagonists & inhibitors; Red Fluorescent Protein; Schwann Cells - cytology; Targeted therapeutics; Transduction, Genetic; Tumor Cells, Cultured
• ISSN: 0014-4886; 1090-2430
• DOI: 10.1016/j.expneurol.2017.10.012

Plexiform neurofibromas (PNs), which may be present at birth in up to half of children with type 1 neurofibromatosis (NF1), can cause serious loss of function, such as quadriparesis, and can undergo malignant transformation. Surgery is the first line treatment although the invasive nature of these tumors often prevents complete resection. Recent clinical trials have shown promising success for some drugs, notably selumetinib, an inhibitor of MAP kinase kinase (MEK). We have developed three-dimensional (3D) cell culture models of immortalized cells from NF1 PNs and of control Schwann cells (SCs) that we believe mimic more closely the in vivo condition than conventional two-dimensional (2D) cell culture. Our goal is to facilitate pre-clinical identification of potential targeted therapeutics for these tumors. Three drugs, selumetinib (a MEK inhibitor), picropodophyllin (an IGF-1R inhibitor) and LDN-193189 (a BMP2 inhibitor) were tested with dose-response design in both 2D and 3D cultures for their abilities to block net cell growth. Cell lines grown in 3D conditions showed varying degrees of resistance to the inhibitory actions of all three drugs. For example, control SCs became resistant to growth inhibition by selumetinib in 3D culture. LDN-193189 was the most effective drug in 3D cultures, with only slightly reduced potency compared to the 2D cultures. Characterization of these models also demonstrated increased proteolysis of collagen IV in the matrix by the PN driver cells as compared to wild-type SCs. The proteolytic capacity of the PN cells in the model may be a clinically significant property that can be used for testing the ability of drugs to inhibit their invasive phenotype. •3D cell culture models were developed of NF1-mutant plexiform neurofibromas.•Morphologies of tumor and wild-type cell lines were distinctively different in 3D.•Plexiform neurofibroma tumor cells showed increased capacity for matrix proteolysis.•A volumetric assay for cell proliferation in 3D cultures was developed.•Drugs screening was performed under 2D and 3D culture conditions.•3D culture revealed differential sensitivity to selumetinib (MEK inhibitor).