Schwann Cells pp 161-173 | Cite as

Generation and Use of Merlin-Deficient Human Schwann Cells for a High-Throughput Chemical Genomics Screening Assay

  • Alejandra M. Petrilli
  • Cristina Fernández-ValleEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1739)


Schwannomas are benign nerve tumors that occur sporadically in the general population and in those with neurofibromatosis type 2 (NF2), a tumor predisposition genetic disorder. NF2-associated schwannomas and most sporadic schwannomas are caused by inactivating mutations in Schwann cells in the neurofibromatosis type 2 gene (NF2) that encodes the merlin tumor suppressor. Despite their benign nature, schwannomas and especially vestibular schwannomas cause considerable morbidity. The primary available therapies are surgery or radiosurgery which usually lead to loss of function of the compromised nerve. Thus, there is a need for effective chemotherapies. We established an untransformed merlin-deficient human Schwann cell line for use in drug discovery studies for NF2-associated schwannomas. We describe the generation of human Schwann cells (HSCs) with depletion of merlin and their application in high-throughput screening of chemical libraries to identify compounds that decrease their viability. This NF2-HSC model is amenable for use in independent labs and high-throughput screening (HTS) facilities.

Key words

Human Schwann cell line Schwannoma High-throughput viability assay Neurofibromatosis type 2 (NF2) Drug discovery Merlin 



This work was supported in part by a Drug Discovery Initiative award from the Children’s Tumor Foundation and with the assistance from Dr. Layton Smith, Director, Drug Discovery Florida, Sanford Burnham Prebys Medical Discovery Institute.


  1. 1.
    Rouleau GA, Merel P, Lutchman M, Sanson M, Zucman J, Marineau C, Hoang-Xuan K, Demczuk S, Desmaze C, Plougastel B et al (1993) Alteration in a new gene encoding a putative membrane-organizing protein causes neuro-fibromatosis type 2. Nature 363(6429):515–521. CrossRefPubMedGoogle Scholar
  2. 2.
    Trofatter JA, MacCollin MM, Rutter JL, Murrell JR, Duyao MP, Parry DM, Eldridge R, Kley N, Menon AG, Pulaski K et al (1993) A novel moesin-, ezrin-, radixin-like gene is a candidate for the neurofibromatosis 2 tumor suppressor. Cell 75(4):826CrossRefPubMedGoogle Scholar
  3. 3.
    Agnihotri S, Jalali S, Wilson MR, Danesh A, Li M, Klironomos G, Krieger JR, Mansouri A, Khan O, Mamatjan Y, Landon-Brace N, Tung T, Dowar M, Li T, Bruce JP, Burrell KE, Tonge PD, Alamsahebpour A, Krischek B, Agarwalla PK, Bi WL, Dunn IF, Beroukhim R, Fehlings MG, Bril V, Pagnotta SM, Iavarone A, Pugh TJ, Aldape KD, Zadeh G (2016) The genomic landscape of schwannoma. Nat Genet 48(11):1339–1348. CrossRefPubMedGoogle Scholar
  4. 4.
    Seizinger BR, Martuza RL, Gusella JF (1986) Loss of genes on chromosome 22 in tumorigenesis of human acoustic neuroma. Nature 322(6080):644–647. CrossRefPubMedGoogle Scholar
  5. 5.
    Petrilli AM, Fernandez-Valle C (2016) Role of Merlin/NF2 inactivation in tumor biology. Oncogene 35(5):537–548. CrossRefPubMedGoogle Scholar
  6. 6.
    Blakeley JO, Plotkin SR (2016) Therapeutic advances for the tumors associated with neurofibromatosis type 1, type 2, and schwannomatosis. Neuro-Oncology 18(5):624–638. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Evans DG (2009) Neurofibromatosis type 2 (NF2): a clinical and molecular review. Orphanet J Rare Dis 4:16. CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Hayflick L (1965) The limited in vitro lifetime of human diploid cell strains. Exp Cell Res 37:614–636CrossRefPubMedGoogle Scholar
  9. 9.
    Nair S, Leung H, Collins A, Ramsden R, Wilson J (2007) Primary cultures of human vestibular schwannoma: selective growth of schwannoma cells. Otol Neurotol 28(2):258–263. CrossRefPubMedGoogle Scholar
  10. 10.
    Rutkowski JL, Kirk CJ, Lerner MA, Tennekoon GI (1995) Purification and expansion of human Schwann cells in vitro. Nat Med 1(1):80–83CrossRefPubMedGoogle Scholar
  11. 11.
    Turnbull VJ (2005) Culturing human Schwann cells. Methods Mol Med 107:173–182PubMedGoogle Scholar
  12. 12.
    Keng VW, Watson AL, Rahrmann EP, Li H, Tschida BR, Moriarity BS, Choi K, Rizvi TA, Collins MH, Wallace MR, Ratner N, Largaespada DA (2012) Conditional inactivation of Pten with EGFR overexpression in Schwann cells models sporadic MPNST. Sarcoma 2012:620834. CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Lehmann HC, Chen W, Mi R, Wang S, Liu Y, Rao M, Hoke A (2012) Human Schwann cells retain essential phenotype characteristics after immortalization. Stem Cells Dev 21(3):423–431. CrossRefPubMedGoogle Scholar
  14. 14.
    Hung G, Li X, Faudoa R, Xeu Z, Kluwe L, Rhim JS, Slattery W, Lim D (2002) Establishment and characterization of a schwannoma cell line from a patient with neurofibromatosis 2. Int J Oncol 20(3):475–482PubMedGoogle Scholar
  15. 15.
    Lepont P, Stickney JT, Foster LA, Meng JJ, Hennigan RF, Ip W (2008) Point mutation in the NF2 gene of HEI-193 human schwannoma cells results in the expression of a merlin isoform with attenuated growth suppressive activity. Mutat Res 637(1–2):142–151. CrossRefPubMedGoogle Scholar
  16. 16.
    Sainio M, Jaaskelainen J, Pihlaja H, Carpen O (2000) Mild familial neurofibromatosis 2 associates with expression of merlin with altered COOH-terminus. Neurology 54(5):1132–1138CrossRefPubMedGoogle Scholar
  17. 17.
    Prabhakar S, Messerli SM, Stemmer-Rachamimov AO, Liu TC, Rabkin S, Martuza R, Breakefield XO (2007) Treatment of implantable NF2 schwannoma tumor models with oncolytic herpes simplex virus G47Delta. Cancer Gene Ther 14(5):460–467. CrossRefPubMedGoogle Scholar
  18. 18.
    James MF, Lelke JM, Maccollin M, Plotkin SR, Stemmer-Rachamimov AO, Ramesh V, Gusella JF (2008) Modeling NF2 with human arachnoidal and meningioma cell culture systems: NF2 silencing reflects the benign character of tumor growth. Neurobiol Dis 29(2):278–292. CrossRefPubMedGoogle Scholar
  19. 19.
    Shalem O, Sanjana NE, Hartenian E, Shi X, Scott DA, Mikkelsen TS, Heckl D, Ebert BL, Root DE, Doench JG, Zhang F (2014) Genome-scale CRISPR-Cas9 knockout screening in human cells. Science 343(6166):84–87. CrossRefPubMedGoogle Scholar
  20. 20.
    Petrilli AM, Fuse MA, Donnan MS, Bott M, Sparrow NA, Tondera D, Huffziger J, Frenzel C, Malany CS, Echeverri CJ, Smith L, Fernandez-Valle C (2014) A chemical biology approach identified PI3K as a potential therapeutic target for neurofibromatosis type 2. Am J Transl Res 6(5):471–493PubMedPubMedCentralGoogle Scholar
  21. 21.
    Zhang JH, Chung TD, Oldenburg KR (1999) A simple statistical parameter for use in evaluation and validation of high throughput screening assays. J Biomol Screen 4(2):67–73CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  • Alejandra M. Petrilli
    • 1
  • Cristina Fernández-Valle
    • 1
    Email author
  1. 1.Neuroscience Division, Burnett School of Biomedical Science, College of MedicineUniversity of Central FloridaOrlandoUSA

Personalised recommendations