CSIG-24. TARGETED INHIBITION OF CDK5-MEDIATED REGULATION OF HUMAN ENDOGENOUS RETROVIRUS K (HML-2) ENVELOPE PROTEIN IN ATYPICAL TERATOID RHABDOID TUMOR

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi38-vi38
Author(s):  
Tara Doucet-O'Hare ◽  
Jared Rosenblum ◽  
Brianna DiSanza ◽  
Catherine DeMarino ◽  
Abigail Atkinson ◽  
...  
Keyword(s):  
Cell Viability ◽  
Cell Lines ◽  
Envelope Protein ◽  
Phosphorylation Site ◽  
Endogenous Retrovirus ◽  
Rhabdoid Tumor ◽  
Human Endogenous Retrovirus ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Atypical Teratoid ◽  
Targeted Inhibition

Abstract Atypical teratoid rhabdoid tumor (ATRT) is a pediatric brain tumor with a high mortality rate characterized by mutations in/ deletions of SWI/SNF matrix-associated actin-dependent regulator of chromatin sub-family B member 1 (SMARCB1). We previously showed that loss of SMARCB1 causes up-regulation and release of HML-2 subfamily of human endogenous retrovirus K envelope protein (HML-2 ENV), resulting in maintenance of pluripotency. Here, we investigated intracellular trafficking and release of HML-2 ENV. Further, we demonstrate two potential therapeutic strategies to decrease intracellular HML-2 ENV: 1) inhibition of calcium influx by ouabain, a cardiac glycoside toxic to neural stem cells, and 2) targeted inhibition of cyclin-dependent kinase 5 (CDK5), which is restricted to neurons by p35, its activator protein, by TP5. ATRT cell lines and tumor tissue obtained from patients were confirmed for SMARCB1 loss and increased HML-2 ENV. Cell viability and intracellular HML-2 ENV concentration were measured after treatment with ouabain and TP5 (CDK5 antagonist). We evaluated the calcium-mediated effect of ouabain on HML-2 intracellular concentration by treating the cells with ouabain, the calcium chelators calcimycin and EGTA, and calpeptin, a calpain inhibitor, which activates CDK5, and measuring HML-2 ENV and p35. We evaluated HML-2 ENV for a CDK5 consensus phosphorylation site and performed co-immunoprecipitation to evaluate direct interaction. We evaluated activity of CDK5 in ATRT cell lines by autoradiography. Both Ouabain and TP5 caused a decrease in cell viability in a dose-dependent manner. Further, ouabain treatment decreased HML-2 ENV intracellular concentration. We found that HML-2 ENV contains a consensus phosphorylation site for CDK5. We discovered that HML-2 ENV was bound to CDK5. We established that ATRT cell lines had hyperactive CDK5. Finally, we established that the effect of ouabain on HML-2 ENV was due to indirect inhibition of calcium-mediated activation of calpain and thus CDK5.

scholarly journals ATRT-07. DEFINING LOST AND GAINED TRANSCRIPTIONAL REGULATORY NETWORKS IN ATYPICAL TERATOID RHABDOID TUMOR

2021 ◽  
Vol 23 (Supplement_1) ◽  
pp. i2-i2
Author(s):  
Cody Nesvick ◽  
Liang Zhang ◽  
Alex Wixom ◽  
Feda Hamdan ◽  
Steven Johnsen ◽  
...  
Keyword(s):  
Cell Lines ◽  
Regulatory Networks ◽  
Regulatory Elements ◽  
Rhabdoid Tumor ◽  
Allelic Loss ◽  
Global Changes ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Transcriptional Regulatory Networks ◽  
Transcriptional Regulatory ◽  
Atypical Teratoid

Abstract Atypical teratoid rhabdoid tumor (ATRT) is a central nervous system cancer of infancy and early childhood that may occur anywhere along the neuraxis and is associated with a high rate of mortality. While contemporary multimodal therapeutic approaches have significantly improved overall survival, targeted therapy remains elusive, and treatment is often associated with significant morbidity. ATRT is unique in its genomic stability, with the only recurrent genetic abnormality being bi-allelic loss of the SMARCB1 gene, which encodes a core subunit of the BAF chromatin remodeling complex. The epigenetic mechanisms by which SMARCB1 loss leads to tumorigenesis are not yet well-defined and addressing this gap in understanding is necessary for creating efficacious, targeted therapeutics. To better understand the epigenetic features gained and lost in ATRT, we re-expressed SMARCB1 in a library of patient-derived and established ATRT cell lines of multiple molecular subtypes. SMARCB1 restoration significantly reduced or eliminated the proliferative and clonogenic capacity of each cell line. We performed assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-Seq) and RNA sequencing (RNA-Seq) to define putative transcriptional regulatory networks that are gained and lost in ATRT. SMARCB1 restoration was associated with global changes in chromatin openness consistent with the creation of new regulatory elements throughout the genome, and these were associated with induction of a diverse developmental transcriptional signature. Motif enrichment analysis of regions with increased accessibility defined a small but consistent number of centrally enriched transcription factor motifs across cell lines indicative of putative pioneer factors whose functions may be lost in ATRT. Pertinent chromatin immunoprecipitation with sequencing (ChIP-Seq) data will be discussed in the context of lost and gained transcriptional regulatory networks in ATRT and normal cellular development.

scholarly journals SMARCB1 deletion in atypical teratoid rhabdoid tumors results in human endogenous retrovirus K (HML-2) expression

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Tara T. Doucet-O’Hare ◽  
Brianna L. DiSanza ◽  
Catherine DeMarino ◽  
Abigail L. Atkinson ◽  
Jared S. Rosenblum ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cellular Proliferation ◽  
Suppressor Gene ◽  
Endogenous Retrovirus ◽  
Human Endogenous Retrovirus ◽  
Knock Down ◽  
Central Nervous System Cancer ◽  
Atypical Teratoid ◽  
Rhabdoid Tumors

AbstractAtypical Teratoid Rhabdoid Tumor (AT/RT) is a rare pediatric central nervous system cancer often characterized by deletion or mutation of SMARCB1, a tumor suppressor gene. In this study, we found that SMARCB1 regulates Human Endogenous Retrovirus K (HERV-K, subtype HML-2) expression. HML-2 is a repetitive element scattered throughout the human genome, encoding several intact viral proteins that have been associated with stem cell maintenance and tumorigenesis. We found HML-2 env expression in both the intracellular and extracellular compartments in all AT/RT cell lines (n = 4) and in 95% of AT/RT patient tissues (n = 37) evaluated. SMARCB1 knock-down in neural stem cells (NSCs) led to an upregulation of HML-2 transcription. We found that SMARCB1 binds adjacent to the HML-2 promoter, repressing its transcription via chromatin immunoprecipitation; restoration of SMARCB1 expression in AT/RT cell lines significantly downregulated HML-2 expression. Further, targeted downregulation of HML-2 transcription via CRISPR-dCas9 coupled with suppressor proteins led to cellular dispersion, decreased proliferation, and cell death in vitro. HML-2 knock-down with shRNA, siRNA, and CRISPR-dCas9 significantly decreased Ras expression as measured by qRT-PCR, suggesting that HML-2 modulates MAPK/ERK signaling in AT/RT cells. Overexpression of NRAS was sufficient to restore cellular proliferation, and MYC, a transcription factor downstream of NRAS, was bound to the HERV-K LTR significantly more in the absence of SMARCB1 expression in AT/RT cells. We show a mechanism by which these undifferentiated tumors remain pluripotent, and we demonstrate that their formation is aided by aberrant HML-2 activation, which is dependent on SMARCB1 and its interaction with MYC.

scholarly journals MODL-27. MEK INHIBITION WITH TRAMETINIB SLOWS PROGRESSION OF MEDULLOBLASTOMA AND ATYPICAL TERATOID RHABDOID TUMOR IN ORTHOTOPIC XENOGRAFT MURINE MODEL

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii416-iii416
Author(s):  
Sujata Mushrif ◽  
Long Hung ◽  
Sakunthala Muthugounder ◽  
Shahab Asgharzadeh
Keyword(s):  
Cell Lines ◽  
Murine Model ◽  
Mapk Pathway ◽  
Autologous Transplantation ◽  
Rhabdoid Tumor ◽  
Clinical Samples ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Cell Viability Assay ◽  
Murine Tumors ◽  
Atypical Teratoid

Abstract BACKGROUND Combination of surgery, chemotherapy, autologous transplantation, irradiation constitutes treatment of CNS embryonal-cell tumors (Medulloblastoma-MBL, atypical teratoid rhabdoid tumor-AT/RT). Targeted agents to improve survival and decrease side effects are necessary. We hypothesize that inhibiting MAPK pathway in MBL and AT/RT may be beneficial. METHODS IHC(pERK) was performed on clinical tumors. Trametinib(MEK inhibitor) was tested on MBL(UW228, D283, DAOY); AT/RT(CHLA06, BT12) cell-lines. Luminescent cell-viability assay was done(72 hrs) and with crystal violet assay(10 days). Orthotopic, xenografts of MBL and AT/RT were made in NOD-Scid gamma mice. Mice were given Trametinib daily by gavage for 6 weeks(0.6mg/kg b.w). Western blot was performed on protein from cell lines and tumor xenografts incubated with Trametinib. H&E staining was done on murine tumors. RESULTS AT/RT(48%) and MBL(57%); Anaplastic(50%), Desmoplastic(40%), Classic(38%); Group 4(66%), Group 3(20%), SHH(55%), WNT(0%) showed presence of pERK(clinical samples). In-vitro, Trametinib completely abrogated the phosphorylation of ERK at 125nM in AT/RT and 50nM in MBL. The IC50 after 10 days exposure was 10nM for AT/RT and 35nM for MBL. Trametinib treated mice showed delay in tumor growth and significant survival advantage in both AT/RT (p=0.00336) and MBL (p=0.0069). Murine tumors showed decreased proliferation (H&E). CONCLUSION Trametinib decreased cell proliferation, increased survival in our murine model in both MBL and AT/RT. Pre-clinical results indicate benefits in subgroups of AT/RT and MBL with active MAPK pathway.

scholarly journals HERV-K Gag RNA and Protein Levels Are Elevated in Malignant Regions of the Prostate in Males with Prostate Cancer

Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 449
Author(s):  
Simin D. Rezaei ◽  
Joshua A. Hayward ◽  
Sam Norden ◽  
John Pedersen ◽  
John Mills ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Lines ◽  
Endogenous Retrovirus ◽  
Human Endogenous Retrovirus ◽  
Normal Prostate ◽  
Tissue Samples ◽  
Gag Protein ◽  
Protein Levels ◽  
Prostate Epithelial Cells ◽  
Prostate Cancers

Heightened expression of human endogenous retrovirus (HERV) sequences has been associated with a range of malignancies, including prostate cancer, suggesting that they may serve as useful diagnostic or prognostic cancer biomarkers. We analysed the expression of HERV-K (Gag and Env/Np9 regions), HERV-E 4.1 (Pol and Env regions), HERV-H (Pol) and HERV-W (Gag) sequences in prostate cancer cells lines and normal prostate epithelial cells using qRT-PCR. HERV expression was also analysed in matched malignant and benign prostate tissue samples from men with prostate cancer (n = 27, median age 65.2 years (range 47–70)) and compared to prostate cancer-free male controls (n = 11). Prostate cancer epithelial cell lines exhibited a signature of HERV RNA overexpression, with all HERVs analysed, except HERV-E Pol, showing heightened expression in at least two, but more commonly all, cell lines analysed. Analysis of primary prostate material indicated increased expression of HERV-E Pol but decreased expression of HERV-E Env in both malignant and benign regions of the prostate in men with prostate cancer as compared to those without. Expression of HERV-K Gag was significantly higher in malignant regions of the prostate in men with prostate cancer as compared to matched benign regions and prostate cancer-free men (p < 0.001 for both), with 85.2% of prostate cancers donors showing malignancy-associated upregulation of HERV-K Gag RNA. HERV-K Gag protein was detected in 12/18 (66.7%) malignant tissues using immunohistochemistry, but only 1/18 (5.6%) benign tissue sections. Heightened expression of HERV-K Gag RNA and protein appears to be a sensitive and specific biomarker of prostate malignancy in this cohort of men with prostate carcinoma, supporting its potential utility as a non-invasive, adjunct clinical biomarker.

scholarly journals ATRT-03. IDENTIFICATION OF microRNA-BASED PROGNOSTIC BIOMARKERS AND CANDIDATE THERAPEUTIC AGENTS FOR ATYPICAL TERATOID/RHABDOID TUMOR

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii275-iii276
Author(s):  
Yang Zhang ◽  
Jianguo Xu
Keyword(s):  
Target Genes ◽  
Kinase Inhibitor ◽  
Therapeutic Agents ◽  
Rhabdoid Tumor ◽  
Janus Kinase ◽  
Prognostic Biomarkers ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Overlapping Genes ◽  
Hub Genes ◽  
Atypical Teratoid

Abstract BACKGROUND MicroRNA (miRNA) has been found to be involved in development of many malignant pediatric brain tumors, including atypical teratoid/rhabdoid tumor (AT/RT) that is highly aggressive and carries a dismal prognosis. The current study investigated the potential value of miRNAs and pivotal genes associated with AT/RT using bioinformatics analysis, aiming to identify new prognostic biomarkers and candidate drugs for AT/RT patients. METHODS Differentially expressed miRNAs (DEMs) and genes (DEGs) between AT/RT and normal control samples were obtained from GEO database. The target genes of DEMs were predicted via TargetScanHuman7.2 and miRDB, and then intersected with DEGs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses of overlapping genes were conducted, followed by construction of protein-protein interaction network. Hub genes were determined by Cytoscape software, and their prognostic values were evaluated using Kaplan-Meier analysis. Connectivity Map database was used to identify latent therapeutic agents. RESULTS A total of 11 DEMs (hsa-miR-1224-5p, hsa-miR-128-3p, hsa-miR-17-5p, hsa-miR-18b-5p, hsa-miR-29c-5p, hsa-miR-329-3p, hsa-miR-379-5p, hsa-miR-433-3p, hsa-miR-488-5p, hsa-miR-656-3p and hsa-miR-885-5p) were screened. By intersecting 3275 predicted target genes and 925 DEGs, we finally identified 226 overlapping genes that were enriched in pathways in cancer and MAPK signaling pathway. Four hub genes (GRIA2, NRXN1, SLC6A1 and SYT1) were significantly associated with the overall survival of AT/RT patients. Candidate drugs included histone deacetylase inhibitor (givinostat), DNA synthesis inhibitor (floxuridine), cyclin-dependent kinase inhibitor (purvalanol) and janus kinase inhibitor (lestaurtinib). CONCLUSION In summary, this study systematically analyzed AT/RT-related miRNAs and pivotal genes to provide novel prognostic biomarkers and potential therapeutic agents.

Atypical Teratoid Rhabdoid Tumor in a Newborn: Can IVF Be a Risk Factor?

2021 ◽  
pp. 1-5
Author(s):  
Turkay Rzayev ◽  
Kubra Gokce ◽  
Safak Gucyetmez ◽  
Suheyla Bozkurt ◽  
Adnan Dagcinar ◽  
...  
Keyword(s):  
Risk Factor ◽  
Rhabdoid Tumor ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Atypical Teratoid

Epithelioid Sarcoma Arising in a Long-Term Survivor of an Atypical Teratoid/Rhabdoid Tumor in a Patient With Rhabdoid Tumor Predisposition Syndrome

2021 ◽  
pp. 109352662098649
Author(s):  
Tiffany G Baker ◽  
Michael J Lyons ◽  
Lee Leddy ◽  
David M Parham ◽  
Cynthia T Welsh
Keyword(s):  
Epithelioid Sarcoma ◽  
Rhabdoid Tumor ◽  
Malignant Rhabdoid Tumor ◽  
Atypical Teratoid Rhabdoid Tumor ◽  
Diagnostic Challenge ◽  
Patient Demographics ◽  
Immunohistochemical Markers ◽  
Genetic Aberration ◽  
Atypical Teratoid ◽  
Rhabdoid Tumors

Rhabdoid tumor predisposition syndrome (RTPS) is defined as the presence of a SMARCB1 or SMARCA4 genetic aberration in a patient with malignant rhabdoid tumor. Patients with RTPS are more likely to present with synchronous or metachronous rhabdoid tumors. Based on the current state of rhabdoid tumor taxonomy, these diagnoses are based largely on patient demographics, anatomic location of disease, and immunohistochemistry, despite their nearly identical histologic and immunohistochemical profiles. Thus, the true distinction between such tumors remains a diagnostic challenge. Central nervous system atypical teratoid/rhabdoid tumor (AT/RT) is a rare, aggressive, primarily pediatric malignancy with variable histologic features and a well documented association with loss of SMARCB1 expression. Epithelioid sarcoma (ES) is a rare soft tissue tumor arising in patients of all ages and characteristically staining for both mesenchymal and epithelial immunohistochemical markers while usually demonstrating loss of SMARCB1 expression. To our knowledge we herein present the first documented case of a patient with RTPS who presented with metachronous AT/RT and ES.

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