Clinical Predictors of Survival for Patients with Atypical Teratoid/Rhabdoid Tumors

Purpose: Atypical teratoid/rhabdoid tumors (AT/RT) are malignant central nervous system (CNS) neoplasms of the young. Our study analyzed a large AT/RT cohort from the National Cancer Database (NCDB) to elucidate predictors of short-term mortality and overall survival (OS). Methods: Information was collected on patients with histologically-confirmed AT/RT using the NCDB (2004-2016). Kaplan-Meier analysis indicated OS. Prognostic factors for 30-day mortality, 90-day mortality, and OS were determined via multivariate Cox proportional-hazards (CPH) and logistic regression models. Results: Our cohort of 189 patients had a median age of 1 year (IQR [1, 4]) and tumor size of 4.7±2.0 cm at diagnosis. Seventy-two percent were under 3 years old; 55.6% were male and 71.0% were Caucasian. Fifty (27.2%) patients received only surgery (S) (OS=5.91 months), 51 (27.7%) received surgery and chemotherapy (S+CT) (OS=11.2 months), and 9 (4.89%) received surgery and radiotherapy (S+RT) (OS=10.3 months). Forty-five (24.5%) received S+CT+RT combination therapy (OS=45.4 months), 13 (17.1%) received S+CT+BMT/SCT (bone marrow or stem cell transplant) (OS=55.5 months), and 16 (8.70%) received S+CT+RT+BMT/SCT (OS=68.4 months). Bivariate analysis of dichotomized age (HR=0.550, 95% CI[0.357, 0.847], p=0.0067) demonstrated significantly increased patient survival if diagnosed at or above 1 year old. On multivariate analysis, administration of S+CT+RT, S+CT+BMT/SCT, or S+CT+RT+BMT/SCT combination therapy predicted significantly (p<0.05) increased OS compared to surgery alone. Conclusion: AT/RTs are CNS tumors where those diagnosed under 1 year old have a significantly worse prognosis. Our study demonstrates that while traditional CT, RT, and BMT/SCT combination regimens prolong life, overall survival in this population is still low.

EXTH-69. FUNCTIONAL GENOMICS UNCOVER GENETIC DEPENDENCIES IN ATRTS

Brain tumors are the leading cause of cancer-related deaths in children. Embryonal brain tumors including medulloblastoma and atypical teratoid rhabdoid tumors (ATRTs) account for 15% of all primary brain and CNS tumors under the age of 14 years, with ATRTs being most prevalent in infants. Despite intensive research efforts, survival estimates for ATRT patients stay relatively low as compared to other tumor entities with a median survival of around 17 months. We here describe genome-wide CRISPR/Cas9 knockout screens in combination with small-molecule drug assays to identify targetable vulnerabilities in ATRTs. Based on functional genomic screening revealing ATRT context-specific genetic vulnerabilities (n = 671 genes), we successfully generated a small-molecule library that shows preferential activity in ATRT cells as compared to a broad selection of other human cancer cell lines. Of note, none of these drugs differentially affect ATRT cells from distinct molecular subgroups, suggesting that top candidate inhibitors might serve as pan-ATRT therapeutic avenues. CDK4/6 inhibitors, among the most potent drugs in our library, are capable of inhibiting tumor growth due to mutual exclusive dependency of ATRTs on either CDK4 or CDK6. Our approach might serve as a blueprint for fostering the identification of functionally-instructed therapeutic strategies in other incurable diseases beyond ATRT, whose genomic profiles also lack actionable alterations so far.

Case Report: The Emerging Role of Ring Chromosome 22 in Phelan-McDermid Syndrome With Atypical Teratoid/Rhabdoid Tumor: The First Child Treated With Growth Hormone

Atypical teratoid/rhabdoid tumors (AT/RTs) in the rhabdoid tumor predisposition syndromes are most often caused by germline mutations of the SMARCB1 gene located in chromosome 22q11.2. Although rarely, it can also result from the constitutional ring chromosome 22 (r22): during mitosis the ring chromosome may lead to an increased rate of somatic mutations, resulting in rhabdoid tumor predispositions when the tumor-suppressor gene SMARCB1 is involved. Individuals with r22 may present similar features as those with Phelan-McDermid syndrome (PMDS) due to 22q13.3 deletion, including the SHANK3 gene. Despite several reports on AT/RT in children with r22 and/or PMDS have been published, the role of constitutional r22 as new oncogenic mechanism for AT/RT is still under investigation. There is not a lot of data available on therapeutic and prognostic implications of r22 in AT/RT and PMDS. Herein, we present the first case of a child with constitutional r22, PMDS and AT/RT of the brain, who is a long term survivor and is been treated with growth hormone. We also describe an unexpected adverse reaction to midazolam.

Natural and cryptic peptides dominate the immunopeptidome of atypical teratoid rhabdoid tumors

BackgroundAtypical teratoid/rhabdoid tumors (AT/RT) are highly aggressive CNS tumors of infancy and early childhood. Hallmark is the surprisingly simple genome with inactivating mutations or deletions in the SMARCB1 gene as the oncogenic driver. Nevertheless, AT/RTs are infiltrated by immune cells and even clonally expanded T cells. However, it is unclear which epitopes T cells might recognize on AT/RT cells.MethodsHere, we report a comprehensive mass spectrometry (MS)-based analysis of naturally presented human leukocyte antigen (HLA) class I and class II ligands on 23 AT/RTs. MS data were validated by matching with a human proteome dataset and exclusion of peptides that are part of the human benignome. Cryptic peptide ligands were identified using Peptide-PRISM.ResultsComparative HLA ligandome analysis of the HLA ligandome revealed 55 class I and 139 class II tumor-exclusive peptides. No peptide originated from the SMARCB1 region. In addition, 61 HLA class I tumor-exclusive peptide sequences derived from non-canonically translated proteins. Combination of peptides from natural and cryptic class I and class II origin gave optimal representation of tumor cell compartments. Substantial overlap existed with the cryptic immunopeptidome of glioblastomas, but no concordance was found with extracranial tumors. More than 80% of AT/RT exclusive peptides were able to successfully prime CD8+ T cells, whereas naturally occurring memory responses in AT/RT patients could only be detected for class II epitopes. Interestingly, >50% of AT/RT exclusive class II ligands were also recognized by T cells from glioblastoma patients but not from healthy donors.ConclusionsThese findings highlight that AT/RTs, potentially paradigmatic for other pediatric tumors with a low mutational load, present a variety of highly immunogenic HLA class I and class II peptides from canonical as well as non-canonical protein sources. Inclusion of such cryptic peptides into therapeutic vaccines would enable an optimized mapping of the tumor cell surface, thereby reducing the likelihood of immune evasion.

Targeting of cyclin-dependent kinases in atypical teratoid rhabdoid tumors with multikinase inhibitor TG02

OBJECTIVE Atypical teratoid rhabdoid tumors (ATRTs) are aggressive pediatric brain tumors with no current standard of care and an estimated median patient survival of 12 to 18 months. Previous genetic analyses have implicated cyclin D1 and enhancer of zeste homolog 2 (EZH2), a histone methyltransferase that is implicated in many cancers, as key drivers of tumorigenicity in ATRTs. Since the effects of EZH2 and cyclin D1 are facilitated by a host of cyclin-dependent kinases (CDKs), the authors sought to investigate the potential therapeutic effects of targeting CDKs in ATRTs with the multi–CDK inhibitor, TG02. METHODS Human ATRT cell lines BT12, BT37, CHLA05, and CHLA06 were selected for investigation. The effects of TG02 on cell viability, proliferation, clonogenicity, and apoptosis were assessed via Cell Counting Kit-8 assays, cell counting, clonogenic assays, and flow cytometry, respectively. Similar methods were used to determine the effects of TG02 combined with radiation therapy (RT) or cisplatin. Synergism indices for TG02-cisplatin combination therapy were calculated using CompuSyn software. RESULTS TG02 was observed to significantly impair ATRT cell growth in vitro by limiting cell proliferation and clonogenicity, and by inducing apoptosis. TG02 inhibited ATRT cell proliferation and decreased cell viability in a dose-dependent manner with nanomolar half maximal effective concentration (EC50) values (BT12, 207.0 nM; BT37, 127.8 nM; CHLA05, 29.7 nM; CHLA06, 18.7 nM). TG02 (150 nM) dramatically increased the proportion of apoptotic ATRT cells 72 hours posttreatment (TG02 8.50% vs control 1.52% apoptotic cells in BT12, p < 0.0001; TG02 70.07% vs control 15.36%, p < 0.0001). Combination therapy studies revealed that TG02 acted as a potent radiosensitizer in ATRT cells (BT12 surviving fraction, RT 51.2% vs RT + TG02 21.7%). Finally, CompuSyn analysis demonstrated that TG02 acted synergistically with cisplatin against ATRT cells at virtually all therapeutic doses. These findings were consistent in cell lines that cover all three molecular subgroups of ATRTs. CONCLUSIONS The results of this investigation have established that TG02 is an effective therapeutic against ATRTs in vitro. Given the lack of standard therapy for ATRTs, these findings help fill an unmet need and support further study of TG02 as a potential therapeutic option for patients with this deadly disease.

NATURAL AND CRYPTIC PEPTIDES DOMINATE THE IMMUNOPEPTIDOME OF ATYPICAL TERATOID RHABDOID TUMORS

Atypical teratoid/rhabdoid tumors (AT/RT) are highly aggressive CNS-tumors of infancy and early childhood. Hallmark is the surprisingly simple genome with inactivating mutations or deletions in the SMARCB1 gene as the oncogenic driver. Nevertheless, AT/RTs are infiltrated by immune cells and even clonally expanded T cells. However, it is unclear, which epitopes T-cells might recognize on AT/RT cells. Here, we report a comprehensive MS-based analysis of naturally presented HLA-class-I and class-II ligands on 23 AT/RTs. Comparative HLA ligandome analysis of the HLA-ligandome revealed 55 class-I and 139 class-II tumor-exclusive peptides. No peptide originated from the SMARCB1-region. In addition, 61 HLA-class I tumor-exclusive peptide sequences derived from non-canonically translated proteins. Combination of peptides from natural and cryptic class I and class II origin gave optimal representation of tumor cell compartments. Substantial overlap existed with the cryptic immunopeptidome of glioblastomas but no concordance was found with extracranial tumors. More than 80% of AT/RT-exclusive peptides were able to successfully prime CD8+ T-cells, whereas naturally occurring memory responses in AT/RT-patients could only be detected for class-II epitopes. Interestingly, >50% of AT/RT-exclusive class-II ligands were also recognized by T-cells from glioblastoma patients but not from healthy donors. These findings highlight that AT/RTs, potentially paradigmatic for other pediatric tumors with a low mutational load, present a variety of highly immunogenic HLA-class-I and class-II peptides from canonical as well as non-canonical protein sources. Inclusion of such cryptic peptides into therapeutic vaccines would enable an optimized mapping of the tumor cell surface, thereby reducing the likelihood of immune evasion.

ATRT-05. REPURPOSED ANTI-MALARIAL QUINACRINE ACTIVATES P53 AND INHIBITS ATRT TUMORIGENICITY

Atypical teratoid rhabdoid tumors (ATRTs) are fatal pediatric brain tumors that warrant improved therapies urgently. ATRTs are characterized by loss of INI1, a subunit of the SWI/SNF chromatin-remodeling complex. ATRTs grow aggressively despite majority of primary tumors expressing p53, suggesting inactivation of this tumor suppressor pathway. Reactivation of p53 could be a potential therapeutic strategy for inhibiting ATRT growth. Our laboratory specializes in researching mechanisms contributing to ATRT pathogenesis and potential therapies. In line with this, we investigated an anti-malarial drug called quinacrine that has been safely used in children for decades and can induce p53 in renal cell carcinoma. We used 5 patient-derived ATRT cell lines (BT-37, BT-12, CHLA-06, CHLA-266, CHLA-05) for our studies. We show that ATRT cell lines treated with quinacrine for 6 hours show increased expression of p53, suggesting its activation. Treatment of ATRT cell lines with increasing doses of quinacrine for 24 hours showed dose-dependent decrease in cell growth and proliferation (assessed by MTS assay and BrdU incorporation, P&lt;0.05) and increase in apoptotic cell death (CC-3 and cleaved PARP expression). Nude mice harboring flank tumors of ATRT cell lines and treated with quinacrine for 3 weeks showed significant reduction in tumor growth compared to control animals (P&lt;0.05). Since quinacrine is a substrate for the drug-efflux proteins P-gp/BCRP, we used quinacrine in combination with elacridar (Pgp/BCRP inhibitor) in our intracranial xenograft experiments to increase quinacrine’s retention in the brain. Mice harboring intracranial xenografts of ATRT cells showed increased survival when treated with quinacrine and elacridar (median survival 46 days) compared to control animals (median survival 25 days). These results suggest that quinacrine inhibits ATRT growth, partly by activating p53. Our studies are the first to show quinacrine’s effect on ATRTs and our current experiments include further investigation of quinacrine’s mechanism.