More than 60% of supratentorial ependymomas harbor a
(ZR
) gene fusion (formerly
). To study the biology of ZR
, we developed an autochthonous mouse tumor model using
electroporation (IUE) of the ...embryonic mouse brain. Integrative epigenomic and transcriptomic mapping was performed on IUE-driven ZR
tumors by CUT&RUN, chromatin immunoprecipitation sequencing, assay for transposase-accessible chromatin sequencing, and RNA sequencing and compared with human ZR
-driven ependymoma. In addition to direct canonical NFκB pathway activation, ZR
dictates a neoplastic transcriptional program and binds to thousands of unique sites across the genome that are enriched with PLAGL family transcription factor (TF) motifs. ZR
activates gene expression programs through recruitment of transcriptional coactivators (Brd4, Ep300, Cbp, Pol2) that are amenable to pharmacologic inhibition. Downstream ZR
target genes converge on developmental programs marked by PLAGL TF proteins, and activate neoplastic programs enriched in Mapk, focal adhesion, and gene imprinting networks. SIGNIFICANCE: Ependymomas are aggressive brain tumors. Although drivers of supratentorial ependymoma (
- and
-associated gene fusions) have been discovered, their functions remain unclear. Our study investigates the biology of
-driven ependymoma, specifically mechanisms of transcriptional deregulation and direct downstream gene networks that may be leveraged for potential therapeutic testing.
.
Brain tumors are the leading cause of cancer-related death in children. Tazemetostat is an FDA-approved enhancer of zeste homolog (EZH2) inhibitor. To determine its role in difficult-to-treat ...pediatric brain tumors, we examined EZH2 levels in a panel of 22 PDOX models and confirmed EZH2 mRNA over-expression in 9 GBM (34.6 ± 12.7-fold) and 11 medulloblastoma models (6.2 ± 1.7 in group 3, 6.0 ± 2.4 in group 4) accompanied by elevated H3K27me3 expression. Therapeutic efficacy was evaluated in 4 models (1 GBM, 2 medulloblastomas and 1 ATRT) via systematically administered tazemetostat (250 and 400 mg/kg, gavaged, twice daily) alone and in combination with cisplatin (5 mg/kg, i.p., twice) and/or radiation (2 Gy/day × 5 days). Compared with the untreated controls, tazemetostat significantly (Pcorrected < 0.05) prolonged survival times in IC-L1115ATRT (101% at 400 mg/kg) and IC-2305GBM (32% at 250 mg/kg, 45% at 400 mg/kg) in a dose-dependent manner. The addition of tazemetostat with radiation was evaluated in 3 models, with only one IC-1078MB (group 4) showing a substantial, though not statistically significant, prolongation in survival compared to radiation treatment alone. Combining tazemetostat (250 mg/kg) with cisplatin was not superior to cisplatin alone in any model. Analysis of in vivo drug resistance detected predominance of EZH2-negative cells in the remnant PDOX tumors accompanied by decreased H3K27me2 and H3K27me3 expressions. These data supported the use of tazemetostat in a subset of pediatric brain tumors and suggests that EZH2-negative tumor cells may have caused therapy resistance and should be prioritized for the search of new therapeutic targets.
This study confirms the preservation of EZH2 overexpression in 22 patient-derived orthotopic xenograft models of pediatric brain tumors. The authors demonstrate the activity of an FDA-approved EZH2 inhibitor, tazemetostat, alone and in combination with radiation in a subset of the models, and identifies EZH2-negative cells as potential cause of therapy resistance.
Background. Meningiomas constitute one-third of all primary brain tumors. Although typically benign, about 20% of these tumors recur despite surgery and radiation, and may ultimately prove fatal. ...There are currently no effective chemotherapies for meningioma. We, therefore, set out to develop patient-derived orthotopic xenograft (PDOX) mouse models of human meningioma using tumor. Method. Of nine patients, four had World Health Organization (WHO) grade I tumors, five had WHO grade II tumors, and in this second group two patients also had recurrent (WHO grade III) meningioma. We also classified the tumors according to our recently developed molecular classification system (Types A, B, and C, with C being the most aggressive). We transplanted all 11 surgical samples into the skull base of immunodeficient (SCID) mice. Only the primary and recurrent tumor cells from one patient—both molecular Type C, despite being WHO grades II and III, respectively—led to the formation of meningioma in the resulting mouse models. We characterized the xenografts by histopathology and RNA-seq and compared them with the original tumors. We performed an in vitro drug screen using 60 anti-cancer drugs followed by in vivo validation. Results. The PDOX models established from the primary and recurrent tumors from patient K29 (K29P-PDOX and K29R-PDOX, respectively) replicated the histopathology and key gene expression profiles of the original samples. Although these xenografts could not be subtransplanted, the cryopreserved primary tumor cells were able to reliably generate PDOX tumors. Drug screening in K29P and K29R tumor cell lines revealed eight compounds that were active on both tumors, including three histone deacetylase (HDAC) inhibitors. We tested the HDAC inhibitor Panobinostat in K29R-PDOX mice, and it significantly prolonged mouse survival (p < 0.05) by inducing histone H3 acetylation and apoptosis. Conclusion. Meningiomas are not very amenable to PDOX modeling, for reasons that remain unclear. Yet at least some of the most malignant tumors can be modeled, and cryopreserved primary tumor cells can create large panels of tumors that can be used for preclinical drug testing.
•A novel pair of orthotopic PDX models of glioma bearing IDH1-R132H/R132C mutations.•New mutant IDH1i (SY-435) with standard therapy led to strong therapeutic efficacy.•H3K4/K9 ...methylation/mtDNA-encoded molecules mediate anti-tumor activity of SYC-435.•Discovered MYO1F, CTC1 and BCL9 as novel genes that mediated SYC-435 resistance.
Clinical outcomes in patients with WHO grade II/III astrocytoma, oligodendroglioma or secondary glioblastoma remain poor. Isocitrate dehydrogenase 1 (IDH1) is mutated in > 70% of these tumors, making it an attractive therapeutic target. To determine the efficacy of our newly developed mutant IDH1 inhibitor, SYC-435 (1-hydroxypyridin-2-one), we treated orthotopic glioma xenograft model (IC-BT142AOA) carrying R132H mutation and our newly established orthotopic patient-derived xenograft (PDX) model of recurrent anaplastic oligoastrocytoma (IC-V0914AOA) bearing R132C mutation. In addition to suppressing IDH1 mutant cell proliferation in vitro, SYC-435 (15 mg/kg, daily x 28 days) synergistically prolonged animal survival times with standard therapies (Temozolomide + fractionated radiation) mediated by reduction of H3K4/H3K9 methylation and expression of mitochondrial DNA (mtDNA)-encoded molecules. Furthermore, RNA-seq of the remnant tumors identified genes (MYO1F, CTC1 and BCL9) and pathways (base excision repair, TCA cycle II, sirtuin signaling, protein kinase A, eukaryotic initiation factor 2 and α-adrenergic signaling) as mediators of therapy resistance. Our data demonstrated the efficacy SYC-435 in targeting IDH1 mutant gliomas when combined with standard therapy and identified a novel set of genes that should be prioritized for future studies to overcome SYC-435 resistance.
Pediatric glioblastoma multiforme (pGBM) is a highly aggressive tumor in need of novel therapies. Our objective was to demonstrate the therapeutic efficacy of MLN8237 (alisertib), an orally available ...selective inhibitor of Aurora A kinase (AURKA), and to evaluate which
model system (monolayer or neurosphere) can predict therapeutic efficacy
AURKA mRNA expressions were screened with qRT-PCR.
antitumor effects were examined in three matching pairs of monolayer and neurosphere lines established from patient-derived orthotopic xenograft (PDOX) models of the untreated (IC-4687GBM), recurrent (IC-3752GBM), and terminal (IC-R0315GBM) tumors, and
therapeutic efficacy through log rank analysis of survival times in two models (IC-4687GBM and IC-R0315GBM) following MLN8237 treatment (30 mg/kg/day, orally, 12 days). Drug concentrations
and mechanism of action and resistance were also investigated.
AURKA mRNA overexpression was detected in 14 pGBM tumors, 10 PDOX models, and 6 cultured pGBM lines as compared with 11 low-grade gliomas and normal brains. MLN8237 penetrated into pGBM xenografts in mouse brains. Significant extension of survival times were achieved in IC-4687GBM of which both neurosphere and monolayer were inhibited
, but not in IC-R0315GBM of which only neurosphere cells responded (similar to IC-3752GBM). Apoptosis-mediated MLN8237 induced cell death, and the presence of AURKA-negative and CD133
cells appears to have contributed to
therapy resistance.
MLN8237 successfully targeted AURKA in a subset of pGBMs. Our data suggest that combination therapy should aim at AURKA-negative and/or CD133
pGBM cells to prevent tumor recurrence.
.
To identify cellular and molecular changes that driver pediatric low grade glioma (PLGG) progression, we analyzed putative cancer stem cells (CSCs) and evaluated key biological changes in a novel and ...progressive patient-derived orthotopic xenograft (PDOX) mouse model. Flow cytometric analysis of 22 PLGGs detected CD133
(<1.5%) and CD15
(20.7 ± 28.9%) cells, and direct intra-cranial implantation of 25 PLGGs led to the development of 1 PDOX model from a grade II pleomorphic xanthoastrocytoma (PXA). While CSC levels did not correlate with patient tumor progression, neurosphere formation and
tumorigenicity, the PDOX model, IC-3635PXA, reproduced key histological features of the original tumor. Similar to the patient tumor that progressed and recurred, IC-3635PXA also progressed during serial
subtransplantations (4 passages), exhibiting increased tumor take rate, elevated proliferation, loss of mature glial marker (GFAP), accumulation of GFAP
/Vimentin
cells, enhanced local invasion, distant perivascular migration, and prominent reactive gliosis in normal mouse brains. Molecularly, xenograft cells with homozygous deletion of
shifted from disomy chromosome 9 to trisomy chromosome 9; and
V600E mutation allele frequency increased (from 28% in patient tumor to 67% in passage III xenografts).
drug screening identified 2/7
V600E inhibitors and 2/9
inhibitors that suppressed cell proliferation. In summary, we showed that PLGG tumorigenicity was low despite the presence of putative CSCs, and our data supported GFAP
/Vimentin
cells,
homozygous deletion in trisomy chromosome 9 cells, and
mutation as candidate drivers of tumor progression in the PXA xenografts.
Introduction: Although BTK inhibitors (BTKi) are effective therapeutics in the treatment of B cell malignancies, emerging BTK resistance mutations in chronic lymphocytic leukemia (CLL), as well as ...potential growth-promoting kinase-independent scaffolding function of BTK, present a need for improved or new approaches. NX-5948 is a novel, orally administered small molecule that induces specific BTK protein degradation by the cereblon E3 ligase complex without degradation of other cereblon neo-substrates. Importantly, NX-5948 induces degradation of wild-type and mutant forms of BTK in B-cells Noviski et al. 2023 at sub-nanomolar potencies and exhibits potent tumor growth inhibition in TMD8 xenograft models that contain either wild-type BTK or BTKi-resistant mutations Robbins 2021. Here we provide the first disclosure of preliminary safety and efficacy findings from a Phase 1a trial of NX-5948 in patients with relapsed/refractory B cell malignancies.
Methods: NX-5948-301 is a Phase 1, first-in-human, dose-escalation and cohort-expansion trial evaluating the safety, tolerability, and clinical activity of NX-5948 in relapsed/refractory CLL and various subtypes of non-Hodgkin's lymphoma (NHL). Key eligibility criteria: ≥2 prior lines of therapy; measurable or other evaluable disease per indication-specific response criteria; Eastern Cooperative Oncology Group Performance Status (ECOG PS) 0-1. Phase 1a (dose escalation) evaluates safety and tolerability of NX-5948 via a standard 3+3 dose escalation in patients with relapsed/refractory B cell malignancies. Approximately 110 patients (30 in Phase 1a, 80 in Phase 1b) may be enrolled and treated until confirmed disease progression or unacceptable toxicity. Endpoints include dose-limiting toxicities (DLTs); treatment-emergent adverse events (TEAEs); deaths; changes in safety parameters; objective response rate per disease-specific response criteria. Phase 1b (dose expansion) will include up to four expansion cohorts.
Results: As of June 9, 2023, 14 patients were enrolled in Phase 1a and received NX-5948 at 50 mg (n=7), 100 mg (n=4), or 200 mg (n=3) orally once daily. Median age was 65 (range 46-79) years; female/male ratio 28.6%/71.4%; white 92.9%; ECOG PS 0/1 21.4%/78.6%; primary diagnoses were CLL (n=4), diffuse large B cell lymphoma (DLBCL, n=4), mantle cell lymphoma (MCL, n=3), marginal zone lymphoma (MZL, n=2), and follicular lymphoma (FL, n=1). Median number of prior therapies was 4.5 (range 2-10), which included: for CLL - BTKi (n=4/4) and BCL2 inhibitor (n=3/4); for NHL - BTKi (n=5/10), bispecific antibody (n=3/10), and CAR-T (n=2/10). NX-5948 was well tolerated with no DLTs and no TEAEs resulting in drug discontinuation or dose reduction. In addition, there were no NX-5948-related grade ≥3 TEAEs or related serious adverse events. The most common TEAEs were purpura/contusion (57.1%, all below grade 3), nausea (35.7%), and thrombocytopenia (35.7%). No atrial fibrillation/flutter or hypertension was reported. Median duration of treatment was 2.8 (range 0.5-9.6) months with 9/14 patients remaining on treatment. Current data indicate that NX-5948 exhibits dose-dependent pharmacokinetics (PK) and a half-life of ~24 hours, supporting once daily dosing (Figure a). Rapid, robust and sustained BTK degradation was observed in all patients, regardless of absolute BTK starting level, tumor type, or NX-5948 dose (Figure b). Of three evaluable patients with CLL receiving the lowest dose of 50 mg, early signs of clinical activity were observed including one confirmed partial response (PR; at 8 and 16 weeks) and 2 patients with stable disease (SD; at 8 weeks). Further treatment responses will be reported at the time of presentation.
Summary/conclusion: Current findings in this heavily pre-treated population of patients with CLL and NHL are encouraging and indicate that NX-5948 is safe and well tolerated and has clinical activity, supporting continuation of its development in CLL and NHL. NX-5948 also exhibits dose-proportional PK, resulting in rapid, robust and sustained BTK degradation. Additional data with higher dose levels and longer treatment duration will be presented at the meeting.
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