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Genetic Delivery of RNA Molecules to Alter Expression of EGFR in Brain Cancer Cells
Author(s) -
Sivetz Nicole,
Falotico Sarah C.,
Nekrasov Peter,
Hicks Martin J.
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.910.10
Subject(s) - epidermal growth factor receptor , rna splicing , cancer research , biology , alternative splicing , messenger rna , gene knockdown , rna , epidermal growth factor , gene , cancer , receptor , microbiology and biotechnology , genetics
Glioblastoma multiforme (GBM) is an incurable and aggressive type of brain tumor. It is the most common central nervous system malignancy with a median survival of only 14 months. GBM is characterized by increased activation of one or more types of cell membrane receptors that drive the proliferation of tumor cells. One receptor in particular, epidermal growth factor receptor (EGFR), is dysregulated in about 60% of GBM tumors. The amplification and overexpression of the EGFR gene makes it a primary target for therapy. Although a great deal is known about the biology of EGFR‐activated GBM tumors, therapies against its biologic processes are limited by the blood‐brain barrier. Therapeutic strategies targeting splicing of the pre‐mRNA transcript, before the mRNA is translated into an active peptide, show great promise. In our current approach, we have designed and cloned a therapeutic gene that encodes a pre‐ trans ‐splicing RNA (PTR). Using anti‐sense targeting, the therapeutic PTR delivers an early stop codon and polyadenylation signal into the EGFR pre‐mRNA transcript, leading to a short, non‐membrane bound soluble peptide. To optimize the PTR we are testing alternative antisense binding sequences, as well as distinct splicing elements along the EGFR transcript. We make use of multiple GBM cell lines and RT‐PCR to measure changes in expression of the EGFR transcript. In addition, we are developing a novel approach using a fluorescent PTR (fPTR) to quickly measure changes in expression. Results show that our vectors were expressed at high levels with subsequent changes in EGFR mRNA expression. Our approach to deliver the DNA encoding anti‐EGFR PTRs to alter pre‐mRNA splicing of the EGFR mRNA transcript in GBM cells has the potential to work from the inside‐out, bypassing systemic barriers. Support or Funding Information Monmouth University School of Science Research Program, Bristol‐Myers Squibb, Independent College Fund of New Jersey, Johnson & Johnson