Journal
MOLECULAR THERAPY-NUCLEIC ACIDS
Volume 14, Issue -, Pages 287-300Publisher
CELL PRESS
DOI: 10.1016/j.omtn.2018.12.003
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Funding
- Hackett Postgraduate Research Scholarship from the University of Western Australia
- Comision Nacional de Investigacion Cientifica y Tecnologica (CONICYT) of Chile [CONICYT-PCHA/Doctorado-Nacional/13-21130879, CONICYT-PAI/7813110022]
- School of Medicine at Pontificia Universidad Catolica de Chile
- Novartis Chile
- Australian Research Council [FT130101767]
- Cancer Council of Western Australia Research Fellowship
- National Health and Medical Research Council [APP1165208, APP1069308, APP1147528, APP1130212]
- NIH [R01CA170370, R01DA036906]
- National Breast Cancer Foundation [NC-14-024]
- Vice Rectorate of Investigation at Pontificia Universidad Catolica de Chile
- Australian Research Council [FT130101767] Funding Source: Australian Research Council
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PTEN expression is lost in many cancers, and even small changes in PTEN activity affect susceptibility and prognosis in a range of highly aggressive malignancies, such as melanoma and triple-negative breast cancer (TNBC). Loss of PTEN expression occurs via multiple mechanisms, including mutation, transcriptional repression and epigenetic silencing. Transcriptional repression of PTEN contributes to resistance to inhibitors used in the clinic, such as B-Raf inhibitors in BRAF mutant melanoma. We aimed to activate PTEN expression using the CRISPR system, specifically dead (d) Cas9 fused to the transactivator VP64-p65-Rta (VPR). dCas9-VPR was directed to the PTEN proximal promoter by single-guide RNAs (sgRNAs), in cancer cells that exhibited low levels of PTEN expression. The dCas9-VPR system increased PTEN expression in melanoma and TNBC cell lines, without transcriptional regulation at predicted off-target sgRNA binding sites. PTEN activation significantly repressed downstream oncogenic pathways, including AKT, mTOR, and MAPK signaling. BRAF V600E mutant melanoma cells transduced with dCas9-VPR displayed reduced migration, as well as diminished colony formation in the presence of B-Raf inhibitors, PI3K/mTOR inhibitors, and with combined PI3K/mTOR and B-Raf inhibition. CRISPR-mediated targeted activation of PTEN may provide an alternative therapeutic approach for highly aggressive cancers that are refractory to current treatments.
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