Diverse genetic, epigenetic, and developmental programs drive glioblastoma, an incurable and poorly understood tumor, but their precise characterization remains challenging. Here, we use an integrative approach spanning single-cell RNA-sequencing of 28 tumors, bulk genetic and expression analysis of 401 specimens from the The Cancer Genome Atlas (TCGA), functional approaches, and single-cell lineage tracing to derive a unified model of cellular states and genetic diversity in glioblastoma. We find that malignant cells in glioblastoma exist in four main cellular states that recapitulate distinct neural cell types, are influenced by the tumor microenvironment, and exhibit plasticity. The relative frequency of cells in each state varies between glioblastoma samples and is influenced by copy number amplifications of the CDK4, EGFR, and PDGFRA loci and by mutations in the NF1 locus, which each favor a defined state. Our work provides a blueprint for glioblastoma, integrating the malignant cell programs, their plasticity, and their modulation by genetic drivers.

Gliomas with histone H3 lysine27-to-methionine mutations (H3K27M-glioma) arise primarily in the midline of the central nervous system of young children, suggesting a cooperation between genetics and cellular context in tumorigenesis. Although the genetics of H3K27M-glioma are well characterized, their cellular architecture remains uncharted. We performed single-cell RNA sequencing in 3321 cells from six primary H3K27M-glioma and matched models. We found that H3K27M-glioma primarily contain cells that resemble oligodendrocyte precursor cells (OPC-like), whereas more differentiated malignant cells are a minority. OPC-like cells exhibit greater proliferation and tumor-propagating potential than their more differentiated counterparts and are at least in part sustained by signaling. Our study characterizes oncogenic and developmental programs in H3K27M-glioma at single-cell resolution and across genetic subclones, suggesting potential therapeutic targets in this disease.

Summary Acquired resistance to Docetaxel precedes fatality in hormone-refractory prostate cancer (HRPC). However, strategies that target Docetaxel resistant cells remain elusive. Using in vitro and in vivo models, we identified a subpopulation of cells that survive Docetaxel exposure. This subpopulation lacks differentiation markers and HLA class I (HLAI) antigens, while overexpressing the Notch and Hedgehog signaling pathways. These cells were found in prostate cancer tissues and were related to tumor aggressiveness and poor patient prognosis. Notably, targeting Notch and Hedgehog signaling depleted this population through inhibition of the survival molecules AKT and Bcl-2, suggesting a therapeutic strategy for abrogating Docetaxel resistance in HRPC. Finally, these cells exhibited potent tumor-initiating capacity, establishing a link between chemotherapy resistance and tumor progression.

Intrahepatic cholangiocarcinoma (iCCA) is a fatal bile duct cancer with dismal prognosis and limited therapeutic options. By performing RNA-and exome-sequencing analyses, we report a novel fusion event, FGFR2-PPHLN1 (16%), and damaging mutations in the ARAF oncogene (11%). Here we demonstrate that the chromosomal translocation t(10;12)(q26;q12) leading to FGFR2-PPHLN1 fusion possesses transforming and oncogenic activity, which is successfully inhibited by a selective FGFR2 inhibitor in vitro. Among the ARAF mutations, N217I and G322S lead to activation of the pathway and N217I shows oncogenic potential in vitro. Screening of a cohort of 107 iCCA patients reveals that FGFR2 fusions represent the most recurrent targetable alteration (45%, 17/107), while they are rarely present in other primary liver tumours (0/100 of hepatocellular carcinoma (HCC); 1/21 of mixed iCCA-HCC). Taken together, around 70% of iCCA patients harbour at least one actionable molecular alteration (FGFR2 fusions, IDH1/2, ARAF, KRAS, BRAF and FGF19) that is amenable for therapeutic targeting.

Highlights d A CRISPR screen for HDAC inhibitor sensitizers in DIPG identifies KDM1A (LSD1) d Corin, a dual inhibitor of LSD1 and HDACs, inhibits DIPG growth in vitro and in vivo d Corin alters histone modifications to promote differentiation and block cell cycle d A Corin-induced gene expression signature correlates with prolonged survival in DIPG

Small cell lung cancer (SCLC) accounts for 15% of lung cancer and is almost always linked to inactivating RB1 and TP53 mutations. SCLC frequently responds, albeit briefly, to chemotherapy. The canonical function of the RB1 gene product, pRB, is to repress the E2F transcription factor family. pRB also plays both E2F-dependent and E2F-independent mitotic roles. We performed a synthetic lethal CRISPR/Cas9 screen in an RB1−/− SCLC cell line that conditionally expresses RB1 to identify dependencies that are caused by pRB loss and discovered that RB1−/− SCLC cell lines are hyperdependent on multiple proteins linked to chromosomal segregation, including Aurora B kinase. Moreover, we show that an Aurora B kinase inhibitor is efficacious in multiple preclinical SCLC models at concentrations that are well tolerated in mice. These results suggest that pRB loss is a predictive biomarker for sensitivity to Aurora B kinase inhibitors in SCLC and perhaps other RB1−/− cancers.

◥Purpose: Dexamethasone, a uniquely potent corticosteroid, is frequently administered to patients with brain tumors to decrease tumor-associated edema, but limited data exist describing how dexamethasone affects the immune system systemically and intratumorally in patients with glioblastoma (GBM), particularly in the context of immunotherapy.Experimental Design: We evaluated the dose-dependent effects of dexamethasone when administered with programmed cell death 1 (PD-1) blockade and/or radiotherapy in immunocompetent C57BL/6 mice with syngeneic GL261 and CT-2A GBM tumors. Clinically, the effect of dexamethasone on survival was evaluated in 181 patients with isocitrate dehydrogenase (IDH) wild-type GBM treated with PD-(L)1 blockade, with adjustment for relevant prognostic factors.Results: Despite the inherent responsiveness of GL261 to immune checkpoint blockade, concurrent dexamethasone administration with anti-PD-1 therapy reduced survival in a dosedependent manner. Concurrent dexamethasone also abrogated survival following anti-PD-1 therapy with or without radiotherapy in immune-resistant CT-2A models. Dexamethasone decreased T-lymphocyte numbers by increasing apoptosis, in addition to decreasing lymphocyte functional capacity. Myeloid and natural killer cell populations were also generally reduced by dexamethasone. Thus, dexamethasone appears to negatively affect both adaptive and innate immune responses. As a clinical correlate, a retrospective analysis of 181 consecutive patients with IDH wild-type GBM treated with PD-(L)1 blockade revealed poorer survival among those on baseline dexamethasone. Upon multivariable adjustment with relevant prognostic factors, baseline dexamethasone administration was the strongest predictor of poor survival [reference, no dexamethasone; <2 mg HR, 2.16; 95% confidence interval (CI), 1.30-3.68; P ¼ 0.003 and ≥2 mg HR, 1.97; 95% CI, 1.23-3.16; P ¼ 0.005].Conclusions: Our preclinical and clinical data indicate that concurrent dexamethasone therapy may be detrimental to immunotherapeutic approaches for patients with GBM.

More than 90% of small cell lung cancers (SCLCs) harbor loss-of-function mutations in the tumor suppressor gene RB1. The canonical function of the RB1 gene product, pRB, is to repress the E2F transcription factor family, but pRB also functions to regulate cellular differentiation in part through its binding to the histone demethylase KDM5A (also known as RBP2 or JARID1A). We show that KDM5A promotes SCLC proliferation and SCLC's neuroendocrine differentiation phenotype in part by sustaining expression of the neuroendocrine transcription factor ASCL1. Mechanistically, we found that KDM5A sustains ASCL1 levels and neuroendocrine differentiation by repressing NOTCH2 and NOTCH target genes. To test the role of KDM5A in SCLC tumorigenesis in vivo, we developed a CRISPR/Cas9-based mouse model of SCLC by delivering an adenovirus (or an adeno-associated virus [AAV]) that expresses Cre recombinase and sgRNAs targeting Rb1, Tp53, and Rbl2 into the lungs of Lox-Stop-Lox Cas9 mice. Coinclusion of a KDM5A sgRNA decreased SCLC tumorigenesis and metastasis, and the SCLCs that formed despite the absence of KDM5A had higher NOTCH activity compared to KDM5A +/+ SCLCs. This work establishes a role for KDM5A in SCLC tumorigenesis and suggests that KDM5 inhibitors should be explored as treatments for SCLC.