Spatial genomic, biochemical and cellular mechanisms underlying meningioma heterogeneity and evolution.

Intratumor heterogeneity underlies cancer evolution and treatment resistance, but targetable mechanisms driving intratumor heterogeneity are poorly understood. Meningiomas are the most common primary intracranial tumors and are resistant to all medical therapies, and high-grade meningiomas have significant intratumor heterogeneity. Here we use spatial approaches to identify genomic, biochemical and cellular mechanisms linking intratumor heterogeneity to the molecular, temporal and spatial evolution of high-grade meningiomas. We show that divergent intratumor gene and protein expression programs distinguish high-grade meningiomas that are otherwise grouped together by current classification systems. Analyses of matched pairs of primary and recurrent meningiomas reveal spatial expansion of subclonal copy number variants associated with treatment resistance. Multiplexed sequential immunofluorescence and deconvolution of meningioma spatial transcriptomes using cell types from single-cell RNA sequencing show decreased immune infiltration, decreased MAPK signaling, increased PI3K-AKT signaling and increased cell proliferation, which are associated with meningioma recurrence. To translate these findings to preclinical models, we use CRISPR interference and lineage tracing approaches to identify combination therapies that target intratumor heterogeneity in meningioma cell co-cultures.

Glioblastoma evolution and heterogeneity from a 3D whole-tumor perspective.

Treatment failure for the lethal brain tumor glioblastoma (GBM) is attributed to intratumoral heterogeneity and tumor evolution. We utilized 3D neuronavigation during surgical resection to acquire samples representing the whole tumor mapped by 3D spatial coordinates. Integrative tissue and single-cell analysis revealed sources of genomic, epigenomic, and microenvironmental intratumoral heterogeneity and their spatial patterning. By distinguishing tumor-wide molecular features from those with regional specificity, we inferred GBM evolutionary trajectories from neurodevelopmental lineage origins and initiating events such as chromothripsis to emergence of genetic subclones and spatially restricted activation of differential tumor and microenvironmental programs in the core, periphery, and contrast-enhancing regions. Our work depicts GBM evolution and heterogeneity from a 3D whole-tumor perspective, highlights potential therapeutic targets that might circumvent heterogeneity-related failures, and establishes an interactive platform enabling 360° visualization and analysis of 3D spatial patterns for user-selected genes, programs, and other features across whole GBM tumors.

Everolimus for Children With Recurrent or Progressive Low-Grade Glioma: Results From the Phase II PNOC001 Trial.

PURPOSE: The PNOC001 phase II single-arm trial sought to estimate progression-free survival (PFS) associated with everolimus therapy for progressive/recurrent pediatric low-grade glioma (pLGG) on the basis of phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway activation as measured by phosphorylated-ribosomal protein S6 and to identify prognostic and predictive biomarkers. PATIENTS AND METHODS: Patients, age 3-21 years, with progressive/recurrent pLGG received everolimus orally, 5 mg/m2 once daily. Frequency of driver gene alterations was compared among independent pLGG cohorts of newly diagnosed and progressive/recurrent patients. PFS at 6 months (primary end point) and median PFS (secondary end point) were estimated for association with everolimus therapy. RESULTS: Between 2012 and 2019, 65 subjects with progressive/recurrent pLGG (median age, 9.6 years; range, 3.0-19.9; 46% female) were enrolled, with a median follow-up of 57.5 months. The 6-month PFS was 67.4% (95% CI, 60.0 to 80.0) and median PFS was 11.1 months (95% CI, 7.6 to 19.8). Hypertriglyceridemia was the most common grade ≥3 adverse event. PI3K/AKT/mTOR pathway activation did not correlate with clinical outcomes (6-month PFS, active 68.4% v nonactive 63.3%; median PFS, active 11.2 months v nonactive 11.1 months; P = .80). Rare/novel KIAA1549::BRAF fusion breakpoints were most frequent in supratentorial midline pilocytic astrocytomas, in patients with progressive/recurrent disease, and correlated with poor clinical outcomes (median PFS, rare/novel KIAA1549::BRAF fusion breakpoints 6.1 months v common KIAA1549::BRAF fusion breakpoints 16.7 months; P < .05). Multivariate analysis confirmed their independent risk factor status for disease progression in PNOC001 and other, independent cohorts. Additionally, rare pathogenic germline variants in homologous recombination genes were identified in 6.8% of PNOC001 patients. CONCLUSION: Everolimus is a well-tolerated therapy for progressive/recurrent pLGGs. Rare/novel KIAA1549::BRAF fusion breakpoints may define biomarkers for progressive disease and should be assessed in future clinical trials.

Whole tumor analysis reveals early origin of the TERT promoter mutation and intercellular heterogeneity in TERT expression.

BACKGROUND: The TERT promoter mutation (TPM) is acquired in most IDH-wildtype glioblastomas (GBM) and IDH-mutant oligodendrogliomas (OD) enabling tumor cell immortality. Previous studies on TPM clonality show conflicting results. This study was performed to determine whether TPM is clonal on a tumor-wide scale. METHODS: We investigated TPM clonality in relation to presumed early events in 19 IDH-wildtype GBM and 10 IDH-mutant OD using 3-dimensional comprehensive tumor sampling. We performed Sanger sequencing on 264 tumor samples and deep amplicon sequencing on 187 tumor samples. We obtained tumor purity and copy number estimates from whole exome sequencing. TERT expression was assessed by RNA-seq and RNAscope. RESULTS: We detected TPM in 100% of tumor samples with quantifiable tumor purity (219 samples). Variant allele frequencies (VAF) of TPM correlate positively with chromosome 10 loss in GBM (R = 0.85), IDH1 mutation in OD (R = 0.87), and with tumor purity (R = 0.91 for GBM; R = 0.90 for OD). In comparison, oncogene amplification was tumor-wide for MDM4- and most EGFR-amplified cases but heterogeneous for MYCN and PDGFRA, and strikingly high in low-purity samples. TPM VAF was moderately correlated with TERT expression (R = 0.52 for GBM; R = 0.65 for OD). TERT expression was detected in a subset of cells, solely in TPM-positive samples, including samples equivocal for tumor. CONCLUSIONS: On a tumor-wide scale, TPM is among the earliest events in glioma evolution. Intercellular heterogeneity of TERT expression, however, suggests dynamic regulation during tumor growth. TERT expression may be a tumor cell-specific biomarker.

"De novo replication repair deficient glioblastoma, IDH-wildtype" is a distinct glioblastoma subtype in adults that may benefit from immune checkpoint blockade.

Glioblastoma is a clinically and molecularly heterogeneous disease, and new predictive biomarkers are needed to identify those patients most likely to respond to specific treatments. Through prospective genomic profiling of 459 consecutive primary treatment-naïve IDH-wildtype glioblastomas in adults, we identified a unique subgroup (2%, 9/459) defined by somatic hypermutation and DNA replication repair deficiency due to biallelic inactivation of a canonical mismatch repair gene. The deleterious mutations in mismatch repair genes were often present in the germline in the heterozygous state with somatic inactivation of the remaining allele, consistent with glioblastomas arising due to underlying Lynch syndrome. A subset of tumors had accompanying proofreading domain mutations in the DNA polymerase POLE and resultant "ultrahypermutation". The median age at diagnosis was 50 years (range 27-78), compared with 63 years for the other 450 patients with conventional glioblastoma (p < 0.01). All tumors had histologic features of the giant cell variant of glioblastoma. They lacked EGFR amplification, lacked combined trisomy of chromosome 7 plus monosomy of chromosome 10, and only rarely had TERT promoter mutation or CDKN2A homozygous deletion, which are hallmarks of conventional IDH-wildtype glioblastoma. Instead, they harbored frequent inactivating mutations in TP53, NF1, PTEN, ATRX, and SETD2 and recurrent activating mutations in PDGFRA. DNA methylation profiling revealed they did not align with known reference adult glioblastoma methylation classes, but instead had unique globally hypomethylated epigenomes and mostly classified as "Diffuse pediatric-type high grade glioma, RTK1 subtype, subclass A". Five patients were treated with immune checkpoint blockade, four of whom survived greater than 3 years. The median overall survival was 36.8 months, compared to 15.5 months for the other 450 patients (p < 0.001). We conclude that "De novo replication repair deficient glioblastoma, IDH-wildtype" represents a biologically distinct subtype in the adult population that may benefit from prospective identification and treatment with immune checkpoint blockade.

Targeted gene expression profiling predicts meningioma outcomes and radiotherapy responses.

Surgery is the mainstay of treatment for meningioma, the most common primary intracranial tumor, but improvements in meningioma risk stratification are needed and indications for postoperative radiotherapy are controversial. Here we develop a targeted gene expression biomarker that predicts meningioma outcomes and radiotherapy responses. Using a discovery cohort of 173 meningiomas, we developed a 34-gene expression risk score and performed clinical and analytical validation of this biomarker on independent meningiomas from 12 institutions across 3 continents (N = 1,856), including 103 meningiomas from a prospective clinical trial. The gene expression biomarker improved discrimination of outcomes compared with all other systems tested (N = 9) in the clinical validation cohort for local recurrence (5-year area under the curve (AUC) 0.81) and overall survival (5-year AUC 0.80). The increase in AUC compared with the standard of care, World Health Organization 2021 grade, was 0.11 for local recurrence (95% confidence interval 0.07 to 0.17, P < 0.001). The gene expression biomarker identified meningiomas benefiting from postoperative radiotherapy (hazard ratio 0.54, 95% confidence interval 0.37 to 0.78, P = 0.0001) and suggested postoperative management could be refined for 29.8% of patients. In sum, our results identify a targeted gene expression biomarker that improves discrimination of meningioma outcomes, including prediction of postoperative radiotherapy responses.

Novel SOX10 indel mutations drive schwannomas through impaired transactivation of myelination gene programs.

BACKGROUND: Schwannomas are common peripheral nerve sheath tumors that can cause severe morbidity given their stereotypic intracranial and paraspinal locations. Similar to many solid tumors, schwannomas and other nerve sheath tumors are primarily thought to arise due to aberrant hyperactivation of the RAS growth factor signaling pathway. Here, we sought to further define the molecular pathogenesis of schwannomas. METHODS: We performed comprehensive genomic profiling on a cohort of 96 human schwannomas, as well as DNA methylation profiling on a subset. Functional studies including RNA sequencing, chromatin immunoprecipitation-DNA sequencing, electrophoretic mobility shift assay, and luciferase reporter assays were performed in a fetal glial cell model following transduction with wildtype and tumor-derived mutant isoforms of SOX10. RESULTS: We identified that nearly one-third of sporadic schwannomas lack alterations in known nerve sheath tumor genes and instead harbor novel recurrent in-frame insertion/deletion mutations in SOX10, which encodes a transcription factor responsible for controlling Schwann cell differentiation and myelination. SOX10 indel mutations were highly enriched in schwannomas arising from nonvestibular cranial nerves (eg facial, trigeminal, vagus) and were absent from vestibular nerve schwannomas driven by NF2 mutation. Functional studies revealed these SOX10 indel mutations have retained DNA binding capacity but impaired transactivation of glial differentiation and myelination gene programs. CONCLUSIONS: We thus speculate that SOX10 indel mutations drive a unique subtype of schwannomas by impeding proper differentiation of immature Schwann cells.

Patterns of Extraneural Metastases in Children With Ependymoma.

Ependymomas account for 10% of all malignant pediatric central nervous system tumors. Standard therapy includes maximal safe surgical resection, followed by focal radiation. Despite the aggressive therapy, progression-free survival is poor. Most ependymoma relapses occur locally at the original tumor site. Extraneural presentations of ependymoma are extremely rare, and no standard of care treatment exists. We present a single-institution case series of 3 patients who experienced extraneural relapses of supratentorial ependymoma and describe their treatment and outcome. These cases of extraneural relapse highlight the possible modes of extraneural spread, including hematogenous, lymphatic, and microscopic seeding through surgical drains and shunts. In addition, they illustrate the increase in histologic grade and mutational burden that may occur at the time of relapse. These cases illustrate the role of aggressive, individualized treatment interventions using a combination of surgery, radiation, and chemotherapy.

Arthritis Severity and Medical Comorbidities Are Prognostic of Worse Outcomes Following Arthroscopic Rotator Cuff Repair in Patients With Concomitant Glenohumeral Osteoarthritis.

Purpose: To assess demographic factors, comorbidities, radiographic variables, and injury patterns as potential prognostic indicators of poor functional and patient-reported outcomes following arthroscopic rotator cuff repair in patients with concomitant glenohumeral osteoarthritis. Methods: A retrospective review of consecutive patients with glenohumeral osteoarthritis who underwent arthroscopic supraspinatus repairs between 2013 and 2018 with a minimum of 1-year follow up was performed. Demographic variables included age, tobacco use, alcohol use, diabetes, sex, hypercholesterolemia, and body mass index while injury patterns included partial- versus full-thickness tear, bicep tendon involvement, and osteoarthritis severity. Multivariate linear regression was used to identify independent predictors of visual analog pain scale (VAS), subjective shoulder value (SSV), and American Shoulder and Elbow Surgeons (ASES) score as well as active range of motion (ROM) in forward flexion (FF) and external rotation (ER). Binary logistic regression was used to identify predictors of repair failure as well as postoperative strength in FF and ER. Results: In total, 91 patients (mean age 61.48 ± 9.4 years) were identified with an average follow up of 26.3 ± 5.7 months. Repair failures occurred in 9.9% (9/91 patients) of the total cohort. Postoperative outcomes were significantly improved with regards to visual analog pain scale, subjective shoulder value, ASES score, ROM in FF, FF strength, and external rotation strength compared with preoperative baseline. Obesity (P = .023) and diabetes (P = .010) were significant independent predictors of greater pain scores postoperatively. Obesity (P = .029) and tobacco use (P = .007) were significant predictors of lower ASES scores postoperatively. Finally, moderate-to-severe osteoarthritis was a significant risk factor for poor ROM and strength in FF postoperatively compared to mild osteoarthritis (P = .029). No variables were predictive of repair failure. Conclusions: Tobacco use, obesity, and diabetes are associated with worse pain and patient-reported outcomes following arthroscopic rotator cuff repair in the context of glenohumeral OA. In addition, moderate-to-severe OA is associated with worse strength and forward flexion compared to those with mild OA. Level of Evidence: Level III, retrospective cohort study.

Conserved features of TERT promoter duplications reveal an activation mechanism that mimics hotspot mutations in cancer.

Mutations in the TERT promoter represent the genetic underpinnings of tumor cell immortality. Beyond the two most common point mutations, which selectively recruit the ETS factor GABP to activate TERT, the significance of other variants is unknown. In seven cancer types, we identify duplications of wildtype sequence within the core promoter region of TERT that have strikingly similar features including an ETS motif, the duplication length and insertion site. The duplications recruit a GABP tetramer by virtue of the native ETS motif and its precisely spaced duplicated counterpart, activate the promoter and are clonal in a TERT expressing multifocal glioblastoma. We conclude that recurrent TERT promoter duplications are functionally and mechanistically equivalent to the hotspot mutations that confer tumor cell immortality. The shared mechanism of these divergent somatic genetic alterations suggests a strong selective pressure for recruitment of the GABP tetramer to activate TERT.

Multiplatform molecular analyses refine classification of gliomas arising in patients with neurofibromatosis type 1.

Gliomas arising in the setting of neurofibromatosis type 1 (NF1) are heterogeneous, occurring from childhood through adulthood, can be histologically low-grade or high-grade, and follow an indolent or aggressive clinical course. Comprehensive profiling of genetic alterations beyond NF1 inactivation and epigenetic classification of these tumors remain limited. Through next-generation sequencing, copy number analysis, and DNA methylation profiling of gliomas from 47 NF1 patients, we identified 2 molecular subgroups of NF1-associated gliomas. The first harbored biallelic NF1 inactivation only, occurred primarily during childhood, followed a more indolent clinical course, and had a unique epigenetic signature for which we propose the terminology "pilocytic astrocytoma, arising in the setting of NF1". The second subgroup harbored additional oncogenic alterations including CDKN2A homozygous deletion and ATRX mutation, occurred primarily during adulthood, followed a more aggressive clinical course, and was epigenetically diverse, with most tumors aligning with either high-grade astrocytoma with piloid features or various subclasses of IDH-wildtype glioblastoma. Several patients were treated with small molecule MEK inhibitors that resulted in stable disease or tumor regression when used as a single agent, but only in the context of those tumors with NF1 inactivation lacking additional oncogenic alterations. Together, these findings highlight recurrently altered pathways in NF1-associated gliomas and help inform targeted therapeutic strategies for this patient population.

EANO - EURACAN - SNO Guidelines on circumscribed astrocytic gliomas, glioneuronal, and neuronal tumors.

In the new WHO 2021 Classification of CNS Tumors the chapter "Circumscribed astrocytic gliomas, glioneuronal and neuronal tumors" encompasses several different rare tumor entities, which occur more frequently in children, adolescents, and young adults. The Task Force has reviewed the evidence of diagnostic and therapeutic interventions, which is low particularly for adult patients, and draw recommendations accordingly. Tumor diagnosis, based on WHO 2021, is primarily performed using conventional histological techniques; however, a molecular workup is important for differential diagnosis, in particular, DNA methylation profiling for the definitive classification of histologically unresolved cases. Molecular factors are increasing of prognostic and predictive importance. MRI finding are non-specific, but for some tumors are characteristic and suggestive. Gross total resection, when feasible, is the most important treatment in terms of prolonging survival and achieving long-term seizure control. Conformal radiotherapy should be considered in grade 3 and incompletely resected grade 2 tumors. In recurrent tumors reoperation and radiotherapy, including stereotactic radiotherapy, can be useful. Targeted therapies may be used in selected patients: BRAF and MEK inhibitors in pilocytic astrocytomas, pleomorphic xanthoastrocytomas, and gangliogliomas when BRAF altered, and mTOR inhibitor everolimus in subependymal giant cells astrocytomas. Sequencing to identify molecular targets is advocated for diagnostic clarification and to direct potential targeted therapies.

Iris and Ciliary Body Melanocytomas Are Defined by Solitary GNAQ Mutation Without Additional Oncogenic Alterations.

OBJECTIVE: To analyze the genetic features of melanocytomas and melanomas of the anterior uvea and assess the value of molecular testing for diagnosis and prognostication. DESIGN: Retrospective case-control study. SUBJECTS: Patients with melanocytoma (n = 16) and melanoma (n = 19) of the anterior uvea. METHODS: Targeted next-generation sequencing was performed on formalin-fixed, paraffin-embedded tumor tissue from anterior uveal melanocytic tumors and correlated with clinicopathologic features. MAIN OUTCOME MEASURES: Presence or absence of accompanying oncogenic alterations beyond GNAQ/GNA11 and their association with histologic features and local recurrence. RESULTS: Hotspot missense mutations in GNAQ/GNA11 were identified in 91% (32/35) of all cases. None of the melanocytomas with or without atypia demonstrated chromosomal imbalances or additional oncogenic variants beyond GNAQ mutation, and none recurred over a median follow-up of 36 months. Additional alterations identified in a subset of melanomas include mutations in BAP1 (n = 3), EIF1AX (n = 4), SRSF2 (n = 1), PTEN (n = 1), and EP300 (n = 1); monosomy 3p (n = 6); trisomy 6p (n = 3); trisomy 8q (n = 2); and an ultraviolet mutational signature (n = 5). Local recurrences were limited to melanomas, all of which demonstrated oncogenic alterations in addition to GNAQ/GNA11 (n = 5). A single melanoma harboring GNAQ and BAP1 mutations and monosomy 3 was the only tumor that metastasized. CONCLUSIONS: In this study, anterior segment uveal melanocytomas did not display oncogenic alterations beyond GNAQ/GNA11. Therefore, they are genetically similar to uveal nevi rather than uveal melanoma based on their molecular features known from the literature. Molecular testing can be performed on borderline cases to aid risk stratification and clinical management decisions.

Targeted Next-Generation Sequencing Reveals Divergent Clonal Evolution in Components of Composite Pleomorphic Xanthoastrocytoma-Ganglioglioma.

Composite pleomorphic xanthoastrocytoma-ganglioglioma (PXA-GG) is an extremely rare central nervous system neoplasm with 2 distinct but intermingled components. Whether this tumor represents a "collision tumor" of separate neoplasms or a monoclonal neoplasm with divergent evolution is poorly understood. Clinicopathologic studies and capture-based next generation sequencing were performed on extracted DNA from all available PXA-GG at 2 medical centers. Five PXA-GG were diagnosed in 1 male and 4 female patients ranging from 13 to 25 years in age. Four arose within the cerebral hemispheres; 1 presented in the cerebellar vermis. DNA was sufficient for analysis in 4 PXA components and 3 GG components. Four paired PXA and GG components harbored BRAF p.V600E hotspot mutations. The 4 sequenced PXA components demonstrated CDKN2A homozygous deletion by sequencing with loss of p16 (protein product of CDKN2A) expression by immunohistochemistry, which was intact in all assessed GG components. The PXA components also demonstrated more frequent copy number alterations relative to paired GG components. In one PXA-GG, shared chromosomal copy number alterations were identified in both components. Our findings support divergent evolution of the PXA and GG components from a common BRAF p.V600E-mutant precursor lesion, with additional acquisition of CDKN2A homozygous deletion in the PXA component as is typically seen in conventional PXA.

Combining radiomics and deep convolutional neural network features from preoperative MRI for predicting clinically relevant genetic biomarkers in glioblastoma.

Background: Glioblastoma is the most common primary brain malignancy, yet treatment options are limited, and prognosis remains guarded. Individualized tumor genetic assessment has become important for accurate prognosis and for guiding emerging targeted therapies. However, challenges remain for widespread tumor genetic testing due to costs and the need for tissue sampling. The aim of this study is to evaluate a novel artificial intelligence method for predicting clinically relevant genetic biomarkers from preoperative brain MRI in patients with glioblastoma. Methods: We retrospectively analyzed preoperative MRI data from 400 patients with glioblastoma, IDH-wildtype or WHO grade 4 astrocytoma, IDH mutant who underwent resection and genetic testing. Nine genetic biomarkers were assessed: hotspot mutations of IDH1 or TERT promoter, pathogenic mutations of TP53, PTEN, ATRX, or CDKN2A/B, MGMT promoter methylation, EGFR amplification, and combined aneuploidy of chromosomes 7 and 10. Models were developed to predict biomarker status from MRI data using radiomics features, convolutional neural network (CNN) features, and a combination of both. Results: Combined model performance was good for IDH1 and TERT promoter hotspot mutations, pathogenic mutations of ATRX and CDKN2A/B, and combined aneuploidy of chromosomes 7 and 10, with receiver operating characteristic area under the curve (ROC AUC) >0.85 and was fair for all other tested biomarkers with ROC AUC >0.7. Combined model performance was statistically superior to individual radiomics and CNN feature models for prediction chromosome 7 and 10 aneuploidy, MGMT promoter methylation, and PTEN mutation. Conclusions: Combining radiomics and CNN features from preoperative MRI yields improved noninvasive genetic biomarker prediction performance in patients with WHO grade 4 diffuse astrocytic gliomas.

CXCL14 Promotes a Robust Brain Tumor-Associated Immune Response in Glioma.

PURPOSE: The immunosuppressive tumor microenvironment present in the majority of diffuse glioma limits therapeutic response to immunotherapy. As the determinants of the glioma-associated immune response are relatively poorly understood, the study of glioma with more robust tumor-associated immune responses may be particularly useful to identify novel immunomodulatory factors that can promote T-cell effector function in glioma. EXPERIMENTAL DESIGN: We used multiplex immune-profiling, proteomic profiling, and gene expression analysis to define the tumor-associated immune response in two molecular subtypes of glioma and identify factors that may modulate this response. We then used patient-derived glioma cultures and an immunocompetent murine model for malignant glioma to analyze the ability of tumor-intrinsic factors to promote a CD8+ T-cell response. RESULTS: As compared with isocitrate dehydrogenase (IDH)-mutant astrocytoma, MAPK-activated pleomorphic xanthoastrocytoma (PXA) harbored increased numbers of activated cytotoxic CD8+ T cells and Iba1+ microglia/macrophages, increased MHC class I expression, enrichment of genes associated with antigen presentation and processing, and increased tumor cell secretion of the chemokine CXCL14. CXCL14 promoted activated CD8+ T-cell chemotaxis in vitro, recruited tumor-infiltrating CD8+ T cells in vivo, and prolonged overall survival in a cytotoxic T-cell-dependent manner. The immunomodulatory molecule B7-H3 was also highly expressed in PXA. CONCLUSIONS: We identify the MAPK-activated lower grade astrocytoma PXA as having an immune-rich tumor microenvironment and suggest this tumor may be particularly vulnerable to immunotherapeutic modulation. We also identify CXCL14 as an important determinant of the glioma-associated immune microenvironment, sufficient to promote an antitumor CD8+ T-cell response.