Erin Eun-Young Ahn, Ph.D., Associate Professor, Molecular and Cellular Pathology, was recently awarded an R01 from the National Cancer Institute based on her project submission, titled, "SON-mediated RNA splicing in glioblastoma." The award began July 1, 2020 and will run through May 30, 2021. The project is expected to run through May 30, 2025.
Dr. Ahn outlined two specific aims for her grant, broken into parts. The first aim is to dissect the molecular mechanisms of SON functions in the regulation of constitutive and alternative RNA splicing in GBM. The second aim is to determine the therapeutic potential of targeting SON in vivo.
The specific goals within the first aim are to determine how SON regulates the U2-associated spliceosome complex during constitutive splicing of PTBP1, define the role of hnRNPs in SON- and RBFOX2-mediated alternative splicing of PTBP2 and the role of RBFOX2 and hnRNP A2B1 in specifying genome-wide SON-RNA interactions in GBM cells, and to conduct a genome-wide analysis of intron retention and alternative splicing upon SON knockdown in patient-derived GSCs. The second aim's goals are to examine the effect of SON knockdown on GBM growth and differentiation in intracranial GSC xenografts and PDX models, examine the effect of combinational targeting of SON and hnRNP A2B1 in intracranial xenograft models, and to examine the therapeutic value of targeting SON together with temozolomide (TMZ) treatment.
Glioblastoma multiforme (GBM) is the most common and lethal brain malignancy with a median survival of only one year after diagnosis. Current knowledge of the underlying basis of GBM centers mostly on several recurrent mutations in specific genes, such as EGFR, p53, IDH1, and PDGFRA. However, non-genetic factors contributing to GBM development and progression are largely unknown. Due to our poor understanding of GBM biology, treatment options are limited to chemotherapy (temozolomide, TMZ) combined with radiotherapy. Thus, new therapeutic approaches are desperately needed to treat this deadly tumor.
RNA splicing is a critical step in determining the amount and the diversity of mRNA, which directly affects protein production. Emerging evidence has demonstrated that aberrant RNA splicing due to splice site mutations and/or splicing factor mutations drives oncogenic gene expression in multiple types of solid tumors and hematologic malignancies. The significance of functional alteration of splicing factors is notable since it can trigger massive and simultaneous changes in a myriad of downstream target RNAs that can switch on the oncogenic gene expression program. In GBM, a few RNA splicing factors, including polypyrimidine tract-binding protein 1 (PTBP1) and hnRNP A2B1 have been recently identified as driving factors in oncogenic splicing, indicating that RNA splicing is a critical, yet to be explored, mechanism that governs a broad range of oncogenic gene expression.
Ahn has been a pioneer in research on SON, a large nuclear speckle protein possessing DNA- and RNA-binding abilities. She previously demonstrated that SON functions as an RNA splicing co-factor and that SON knockdown impairs RNA splicing of a group of genes associated with cell cycle progression and DNA repair/replication. Here, the extensive preliminary data demonstrated that (1) SON is highly upregulated in GBM patient samples and that high SON expression correlates with short patient survival and (2) SON facilitates expression of PTBP1, thereby activating the PTBP1-meditated oncogenic splicing program. In contrast, (3) SON inhibits the expression of PTBP2, a splicing factor required for neural exon inclusion, thereby blocking neural differentiation. We further revealed that (4) SON facilitates the removal of a “detained” intron at the constitutive splicing site in the PTBP1 transcript, and (5) SON inhibits cassette exon (neural exon) inclusion during alternative splicing of PTBP2 in cooperation with hnRNP A2B1 and in competition with RBFOX2 for RNA occupancy. She also demonstrated that (6) SON knockdown markedly inhibits GBM cell growth and reduces the expression of neural stem cell genes. Importantly, we noticed that (7) SON-mediated RNA splicing target genes largely overlap with genes associated with TMZ sensitivity and showed that (8) SON knockdown renders patient-derived glioma stem cells (GSCs) sensitive to TMZ in vitro.
Based on her preliminary data, she hypothesizes that SON is a master RNA splicing regulator positioned at the apex of the splicing factor hierarchy that affects both constitutive and alternative RNA splicing, consequently turning on the oncogenic splicing program and blocking neural splicing. Thus, SON could represent a promising novel therapeutic target for GBM. Her proposal's specified aims will test this hypothesis.
Ahn explains, "We have exceptional expertise to study SON and RNA splicing and have generated extensive preliminary data. We have also recruited the necessary collaborators for our proposed in vivo GBM study and for data analyses."
This proposal is for (1) the comprehensive understanding of underlying molecular mechanisms of SON-mediated RNA splicing in GBM and (2) evaluating the therapeutic value of targeting SON for GBM treatment, thus representing a high level of both biological and clinical significance. Successful completion of this proposed study will significantly advance our knowledge of abnormal gene expression in GBM and provide a fundamental rationale for future endeavors to develop SON inhibitors.
Dr. Ahn joined UAB's Department of Pathology in 2020 as an associate professor. She additionally works as a scientist in the O'Neal Comprehensive Cancer Center. Before joining UAB this year, she was an associate professor of Biochemistry and Molecular Biology and the Mitchell Cancer Center at the University of South Alabama. She earned her M.S. in Cell and Molecular Biology at Seoul National University, South Korea and her Ph.D. in Molecular and Cellular Pathology here at UAB. In 2017, she was named the recipient of the Mayer Mitchell Award for Excellence in Cancer Research. The award is presented annually to a promising scientist at USA Mitchell Cancer Institute upon the recommendation of a faculty committee. In 2015, she was awarded a five-year $1.7 million R01 grant from the National Cancer Institute to study the role of the SON protein in the development of leukemia.
By Hannah Weems