Ashwaq K. Aljabri

Unraveling SMARCA1's Role in Cancer Progression, Drug Resistance, and Muscle Differentiation Through EMT-Related Signaling, the TGF-β Pathway, and Key Transcription Factors SNAI1 and EGR1

Ashwaq K. Aljabri¹² Yuliya Kriga³ Juan Manuel Caravaca³ Jyoti Shetty³ Bao Tran³ Matthew Geisler⁴ Judith K. Davie² Marielle E. Yohe¹ 1. Laboratory of Cell and Developmental Signaling, National Cancer Institute, National Institutes of Health, Frederick, MD, USA 2. Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL, USA 3. Sequencing Facility, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Frederick, MD, USA 4. Department of Plant Biology, Southern Illinois University, Carbondale, IL, USA

SMARCA1, a member of the ISWI family of ATP-dependent chromatin remodelers, plays a crucial role in regulating chromatin structure and gene expression. While its involvement in both normal muscle development and Rhabdomyosarcoma (RMS) has been suggested, no one has yet comprehensively investigated its dual role in these contexts. RMS, a malignant soft tissue sarcoma, is characterized by disrupted myogenic differentiation and aggressive tumor behavior. Data integration of RNA/ATAC-seq analysis of RH30 SMARCA1 knockout (KO) cells, a fusion-positive alveolar Rhabdomyosarcoma (RMS) cell line, revealed significant differential gene expression and chromatin accessibility changes. Top genes such as TGFBR1, MYOG, and SMARCD3, which are essential for muscle differentiation, were highly enriched in RH30 WT compared to SMARCA1 KO cells. Loss of SMARCA1 disrupted chromatin accessibility at these loci, emphasizing its critical role in regulating gene expression for myogenic, growth, and survival pathways. Additionally, EGR1 knockdown in RD cells (fusion-negative embryonal RMS) resulted in the downregulation of MYOD, further demonstrating SMARCA1's influence on transcription factors such as SNAI1, EGR1, PTEN, TBX2, and P53. In SMS-CTR (fusion-negative ERMS) and its resistant derivative, the MEK inhibitor-resistant CTR-557 cell line, RNA and ATAC-seq data showed enrichment of the TGF-β pathway, indicating that this pathway mediates resistance to MEK inhibitors. SMARCA1 interacts with HDAC2, and HDAC2/PI3K inhibition has shown promise in overcoming drug resistance in these models. The findings demonstrate that SMARCA1 is directly enriched on key regulatory genes and transcription factors such as SNAI1, EMC9, PPP2R5C, EGR1, EYA2, SMARCB1, SMARCA4, MYCN, CDK4, and NFKB2, driving important processes such as EMT and cell adhesion, as well as MAPK, WNT, PI3K, and TGF-β pathways. Further, cell survival assays demonstrated that SMARCA1 KO cells exhibited increased sensitivity to the HDAC2 inhibitor Mocetinostat and the PI3K-mTOR inhibitor Dactolisib, suggesting that targeting these pathways could be a promising strategy in SMARCA1-deficient RMS. Targeting both the canonical and non-canonical TGF-β pathways, alongside SMARCA1, offers potential therapeutic strategies to overcome drug resistance and potentially prevent metastasis in resistant cancers.