P/<scp>CAF</scp> mediates <scp>PAX3–FOXO1</scp>‐dependent oncogenesis in alveolar rhabdomyosarcoma

Bharathy, Narendra; Suriyamurthy, Sudha; Rao, Vinay Kumar; Ow, Jin Rong; Lim, Huey Jin; Chakraborty, Payal; Vasudevan, Madavan; Dhamne, Chetan; Chang, Kenneth Tou En; Lee, Victor Kwan Min; Kundu, Tapas K.; Taneja, Reshma

doi:10.1002/path.4773

Cited by 19 publications

(18 citation statements)

References 52 publications

(87 reference statements)

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“…These seemingly opposing effects on protein stability might suggest that protein turnover of WT versus the fusion protein is regulated by different mechanisms, as recently also suggested for the EWS-FLI1 fusion expressed in Ewing sarcoma (48). A similar observation has already been described for acetylation of K426 and K429 by the histone acetyltransferase KAT2B that also stabilize PAX3-FOXO1 (49). Acetylation of the corresponding sites in WT FOXO1 results in phosphorylation at S256 and subsequent degradation (27).…”

Section: Discussionsupporting

confidence: 59%

Aurora A Kinase Inhibition Destabilizes PAX3-FOXO1 and MYCN and Synergizes with Navitoclax to Induce Rhabdomyosarcoma Cell Death

et al. 2020

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◥The clinically aggressive alveolar rhabdomyosarcoma (RMS) subtype is characterized by expression of the oncogenic fusion protein PAX3-FOXO1, which is critical for tumorigenesis and cell survival. Here, we studied the mechanism of cell death induced by loss of PAX3-FOXO1 expression and identified a novel pharmacologic combination therapy that interferes with PAX3-FOXO1 biology at different levels. Depletion of PAX3-FOXO1 in fusion-positive (FP)-RMS cells induced intrinsic apoptosis in a NOXA-dependent manner. This was pharmacologically mimicked by the BH3 mimetic navitoclax, identified as top compound in a screen from 208 targeted compounds. In a parallel approach, and to identify drugs that alter the stability of PAX3-FOXO1 protein, the same drug library was screened and fusion protein levels were directly measured as a read-out. This revealed that inhibition of Aurora kinase A most efficiently negatively affected PAX3-FOXO1 protein levels. Interestingly, this occurred through a novel specific phosphorylation event in and binding to the fusion protein. Aurora kinase A inhibition also destabilized MYCN, which is both a functionally important oncogene and transcriptional target of PAX3-FOXO1. Combined treatment with an Aurora kinase A inhibitor and navitoclax in FP-RMS cell lines and patient-derived xenografts synergistically induced cell death and significantly slowed tumor growth. These studies identify a novel functional interaction of Aurora kinase A with both PAX3-FOXO1 and its effector MYCN, and reveal new opportunities for targeted combination treatment of FP-RMS.Significance: These findings show that Aurora kinase A and Bcl-2 family proteins are potential targets for FP-RMS.

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Section: Discussionsupporting

confidence: 59%

Aurora A Kinase Inhibition Destabilizes PAX3-FOXO1 and MYCN and Synergizes with Navitoclax to Induce Rhabdomyosarcoma Cell Death

et al. 2020

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“…ARMS cell lines RH30, RH4, and RH41 were provided by Peter Houghton (Nationwide Children's Hospital, Columbus, OH) and Rosella Rota (Bambino Gesu Children's Hospital, Rome, Italy) and cultured in RPMI 1640 with L-glutamine (Thermo Fisher Scientific) with 10% FBS (Hyclone). ARMS cells lines were authenticated for PAX3-FOXO1, MyoD, and myogenin expression by Western blot as described (21). Cell lines were tested at least every 6 months for Mycoplasma contamination using the Mycokit Detection Kit from Biowest (K1000).…”

Section: Cell Culture and Stable Cell Linesmentioning

confidence: 99%

“…All animal procedures were approved by the Institutional Animal Care and Use Committee. Note that 6 Â 10 6 RH30 cells were injected subcutaneously into the right flank of 6-week-old CrTac:NCr-Foxn1 nude female mice (InVivos) as described (21). When tumor nodules were palpable, mice (n ¼ 10/group) were injected intraperitoneally with UNC0642 at 5 mg/kg body weight every alternate day.…”

Section: Mouse Xenograft Experiments and Ihcmentioning

confidence: 99%

“…IHC on 15 primary fusion-positive archival tumor tissues obtained from the National University Hospital and KK Women's and Children Hospital in Singapore was done essentially as described (21). Sections were stained with anti-G9a antibody (1:50 dilution, Cell Signaling Technology).…”

Section: Mouse Xenograft Experiments and Ihcmentioning

confidence: 99%

“…Recent advances in chromatin biology have revealed deregulation of the epigenetic landscape in ARMS (18). The chromatin modifiers CHD4, PCAF, and BRD4 have been shown to interact with and affect PAX3-FOXO1 activity (19)(20)(21). Nevertheless, the epigenetic network that is central to ARMS tumorigenesis remains to be identified.…”

Section: Introductionmentioning

confidence: 99%

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Epigenetic Regulation of the PTEN–AKT–RAC1 Axis by G9a Is Critical for Tumor Growth in Alveolar Rhabdomyosarcoma

et al. 2019

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Alveolar rhabdomyosarcoma (ARMS) is an aggressive pediatric cancer with poor prognosis. As transient and stable modifications to chromatin have emerged as critical mechanisms in oncogenic signaling, efforts to target epigenetic modifiers as a therapeutic strategy have accelerated in recent years. To identify chromatin modifiers that sustain tumor growth, we performed an epigenetic screen and found that inhibition of lysine methyltransferase G9a significantly affected the viability of ARMS cell lines. Targeting expression or activity of G9a reduced cellular proliferation and motility in vitro and tumor growth in vivo. Transcriptome and chromatin immunoprecipitation-sequencing analysis provided mechanistic evidence that the tumor-suppressor PTEN was a direct target gene of G9a. G9a repressed PTEN expression in a methyltransferase activitydependent manner, resulting in increased AKT and RAC1 activity. Re-expression of constitutively active RAC1 in G9adeficient tumor cells restored oncogenic phenotypes, demonstrating its critical functions downstream of G9a. Collectively, our study provides evidence for a G9a-dependent epigenetic program that regulates tumor growth and suggests targeting G9a as a therapeutic strategy in ARMS. Significance: These findings demonstrate that RAC1 is an effector of G9a oncogenic functions and highlight the potential of G9a inhibitors in the treatment of ARMS.

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EHMT1 promotes tumor progression and maintains stemness by regulating ALDH1A1 expression in alveolar rhabdomyosarcoma

Nachiyappan

Soon

Lim

et al. 2022

The Journal of Pathology

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Alveolar rhabdomyosarcoma (ARMS) is an aggressive pediatric cancer with poor prognosis. Cancer stem cells (CSCs) are seeds for tumor relapse and metastasis. However, pathways that maintain stemness genes are not fully understood. Here, we report that the enzyme euchromatic histone lysine methyltransferase 1 (EHMT1) is expressed in primary and relapse ARMS tumors. EHMT1 suppression impaired motility and induced differentiation in ARMS cell lines and reduced tumor progression in a mouse xenograft model in vivo. RNA sequencing of EHMT1-depleted cells revealed downregulation of ALDH1A1 that is associated with CSCs. Consistent with this, inhibition of ALDH1A1 expression and activity mimicked EHMT1 depletion phenotypes and reduced tumorsphere formation. Mechanistically, we demonstrate that EHMT1 does not bind to the ALDH1A1 promoter but activates it by stabilizing C/EBPβ, a known regulator of ALDH1A1 expression. Our findings identify a role for EHMT1 in maintenance of stemness by regulating ALDH1A1 expression and suggest that targeting ALDH + cells is a promising strategy in ARMS.

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P/CAF mediates PAX3–FOXO1‐dependent oncogenesis in alveolar rhabdomyosarcoma

Cited by 19 publications

References 52 publications

Aurora A Kinase Inhibition Destabilizes PAX3-FOXO1 and MYCN and Synergizes with Navitoclax to Induce Rhabdomyosarcoma Cell Death

Aurora A Kinase Inhibition Destabilizes PAX3-FOXO1 and MYCN and Synergizes with Navitoclax to Induce Rhabdomyosarcoma Cell Death

Epigenetic Regulation of the PTEN–AKT–RAC1 Axis by G9a Is Critical for Tumor Growth in Alveolar Rhabdomyosarcoma

EHMT1 promotes tumor progression and maintains stemness by regulating ALDH1A1 expression in alveolar rhabdomyosarcoma

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