MicroRNA-142 Is Critical for the Homeostasis and Function of Type 1 Innate Lymphoid Cells.

Natural killer (NK) cells are cytotoxic type 1 innate lymphoid cells (ILCs) that defend against viruses and mediate anti-tumor responses, yet mechanisms controlling their development and function remain incompletely understood. We hypothesized that the abundantly expressed microRNA-142 (miR-142) is a critical regulator of type 1 ILC biology. Interleukin-15 (IL-15) signaling induced miR-142 expression, whereas global and ILC-specific miR-142-deficient mice exhibited a cell-intrinsic loss of NK cells. Death of NK cells resulted from diminished IL-15 receptor signaling within miR-142-deficient mice, likely via reduced suppressor of cytokine signaling-1 (Socs1) regulation by miR-142-5p. ILCs persisting in Mir142-/- mice demonstrated increased expression of the miR-142-3p target αV integrin, which supported their survival. Global miR-142-deficient mice exhibited an expansion of ILC1-like cells concurrent with increased transforming growth factor-ß (TGF-ß) signaling. Further, miR-142-deficient mice had reduced NK-cell-dependent function and increased susceptibility to murine cytomegalovirus (MCMV) infection. Thus, miR-142 critically integrates environmental cues for proper type 1 ILC homeostasis and defense against viral infection.

Acquired copy number alterations of miRNA genes in acute myeloid leukemia are uncommon.

Altered microRNA (miRNA) expression is frequently observed in acute myelogenous leukemia (AML) and has been implicated in leukemic transformation. Whether somatic copy number alterations (CNAs) are a frequent cause of altered miRNA gene expression is largely unknown. Herein, we used comparative genomic hybridization with a custom high-resolution miRNA-centric array and/or whole-genome sequence data to identify somatic CNAs involving miRNA genes in 113 cases of AML, including 50 cases of de novo AML, 18 cases of relapsed AML, 15 cases of secondary AML following myelodysplastic syndrome, and 30 cases of therapy-related AML. We identified a total of 48 somatic miRNA gene-containing CNAs that were not identified by routine cytogenetics in 20 patients (18%). All these CNAs also included one or more protein coding genes. We identified a single case with a hemizygous deletion of MIR223, resulting in the complete loss of miR-223 expression. Three other cases of AML were identified with very low to absent miR-223 expression, an miRNA gene known to play a key role in myelopoiesis. However, in these cases, no somatic genetic alteration of MIR223 was identified, suggesting epigenetic silencing. These data show that somatic CNAs specifically targeting miRNA genes are uncommon in AML.

Complete characterization of the microRNAome in a patient with acute myeloid leukemia.

MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression and have been implicated in the pathogenesis of cancer. In this study, we applied next generation sequencing techniques to comprehensively assess miRNA expression, identify genetic variants of miRNA genes, and screen for alterations in miRNA binding sites in a patient with acute myeloid leukemia. RNA sequencing of leukemic myeloblasts or CD34(+) cells pooled from healthy donors showed that 472 miRNAs were expressed, including 7 novel miRNAs, some of which displayed differential expression. Sequencing of all known miRNA genes revealed several novel germline polymorphisms but no acquired mutations in the leukemia genome. Analysis of the sequence of the 3'-untranslated regions (UTRs) of all coding genes identified a single somatic mutation in the 3'-UTR of TNFAIP2, a known target of the PML-RARα oncogene. This mutation resulted in translational repression of a reporter gene in a Dicer-dependent fashion. This study represents the first complete characterization of the "miRNAome" in a primary human cancer and suggests that generation of miRNA binding sites in the UTR regions of genes is another potential mechanism by which somatic mutations can affect gene expression.

VRK1 as a synthetic lethal target in VRK2 promoter-methylated cancers of the nervous system.

Collateral lethality occurs when loss of a gene/protein renders cancer cells dependent on its remaining paralog. Combining genome-scale CRISPR/Cas9 loss-of-function screens with RNA sequencing in over 900 cancer cell lines, we found that cancers of nervous system lineage, including adult and pediatric gliomas and neuroblastomas, required the nuclear kinase vaccinia-related kinase 1 (VRK1) for their survival in vivo. VRK1 dependency was inversely correlated with expression of its paralog VRK2. VRK2 knockout sensitized cells to VRK1 loss, and conversely, VRK2 overexpression increased cell fitness in the setting of VRK1 loss. DNA methylation of the VRK2 promoter was associated with low VRK2 expression in human neuroblastomas and adult and pediatric gliomas. Mechanistically, depletion of VRK1 reduced barrier-to-autointegration factor phosphorylation during mitosis, resulting in DNA damage and apoptosis. Together, these studies identify VRK1 as a synthetic lethal target in VRK2 promoter-methylated adult and pediatric gliomas and neuroblastomas.

The landscape of tumor cell states and spatial organization in H3-K27M mutant diffuse midline glioma across age and location.

Histone 3 lysine27-to-methionine (H3-K27M) mutations most frequently occur in diffuse midline gliomas (DMGs) of the childhood pons but are also increasingly recognized in adults. Their potential heterogeneity at different ages and midline locations is vastly understudied. Here, through dissecting the single-cell transcriptomic, epigenomic and spatial architectures of a comprehensive cohort of patient H3-K27M DMGs, we delineate how age and anatomical location shape glioma cell-intrinsic and -extrinsic features in light of the shared driver mutation. We show that stem-like oligodendroglial precursor-like cells, present across all clinico-anatomical groups, display varying levels of maturation dependent on location. We reveal a previously underappreciated relationship between mesenchymal cancer cell states and age, linked to age-dependent differences in the immune microenvironment. Further, we resolve the spatial organization of H3-K27M DMG cell populations and identify a mitotic oligodendroglial-lineage niche. Collectively, our study provides a powerful framework for rational modeling and therapeutic interventions.

BAF Complex Maintains Glioma Stem Cells in Pediatric H3K27M Glioma.

Diffuse midline gliomas are uniformly fatal pediatric central nervous system cancers that are refractory to standard-of-care therapeutic modalities. The primary genetic drivers are a set of recurrent amino acid substitutions in genes encoding histone H3 (H3K27M), which are currently undruggable. These H3K27M oncohistones perturb normal chromatin architecture, resulting in an aberrant epigenetic landscape. To interrogate for epigenetic dependencies, we performed a CRISPR screen and show that patient-derived H3K27M-glioma neurospheres are dependent on core components of the mammalian BAF (SWI/SNF) chromatin remodeling complex. The BAF complex maintains glioma stem cells in a cycling, oligodendrocyte precursor cell-like state, in which genetic perturbation of the BAF catalytic subunit SMARCA4 (BRG1), as well as pharmacologic suppression, opposes proliferation, promotes progression of differentiation along the astrocytic lineage, and improves overall survival of patient-derived xenograft models. In summary, we demonstrate that therapeutic inhibition of the BAF complex has translational potential for children with H3K27M gliomas. SIGNIFICANCE: Epigenetic dysregulation is at the core of H3K27M-glioma tumorigenesis. Here, we identify the BRG1-BAF complex as a critical regulator of enhancer and transcription factor landscapes, which maintain H3K27M glioma in their progenitor state, precluding glial differentiation, and establish pharmacologic targeting of the BAF complex as a novel treatment strategy for pediatric H3K27M glioma. See related commentary by Beytagh and Weiss, p. 2730. See related article by Mo et al., p. 2906.

Toddler With New Onset Diabetes and Atypical Hemolytic-Uremic Syndrome in the Setting of COVID-19.

This is a novel case of a 16-month-old boy with a history of prematurity with intrauterine growth restriction, severe failure to thrive, microcephaly, pachygyria, agenesis of the corpus callosum, and postnatal embolic stroke, who presented with new-onset diabetes mellitus with diabetic ketoacidosis in the setting of severe acute respiratory syndrome coronavirus 2 infection, with a course complicated by atypical hemolytic syndrome (aHUS). This patient demonstrated remarkable insulin resistance in the period before aHUS diagnosis, which resolved with the first dose of eculizumab therapy. There is increasing evidence that COVID-19 is associated with thrombotic disorders and that microangiopathic processes and complement-mediated inflammation may be implicated. In this case report, we describe a pediatric patient with COVID-19 and a new complement-mediated microangiopathic thrombotic disease. Because whole-exome sequencing and extensive workup returned without a clear etiology for aHUS, this is likely a COVID-19 triggered case of aHUS versus an idiopathic case that was unmasked by the infection.

Molecular Characterization of Inflammatory Tumors Facilitates Initiation of Effective Therapy.

Inflammatory myofibroblastic tumor (IMT) is a rare, mesenchymal tumor that has an increased incidence in childhood. Tumors are usually isolated to the chest, abdomen, and retroperitoneum, but metastatic presentations can be seen. Presenting symptoms are nonspecific and include fever, weight loss, pain, shortness of breath, and cough. Approximately 85% of IMTs harbor actionable kinase fusions. The diagnosis can be delayed because of overlapping features with inflammatory disorders, such as elevated inflammatory markers, increased immunoglobin G levels, fever, weight loss, and morphologic similarity with nonmalignant conditions. We present a girl aged 11 years with a TFG-ROS1 fusion-positive tumor of the lung that was initially diagnosed as an immunoglobin G4-related inflammatory pseudotumor. She underwent complete left-sided pneumonectomy and later recurred with widely metastatic disease. We then report the case of a boy aged 9 years with widely metastatic TFG-ROS1 fusion-positive IMT with rapid molecular diagnosis. In both children, there was an excellent response to oral targeted therapy. These cases reveal that rapid molecular testing of inflammatory tumors is not only important for diagnosis but also reveals therapeutic opportunities. Targeted inhibitors produce significant radiologic responses, enabling potentially curative treatment approaches for metastatic ROS1 fusion IMT with previously limited treatment options. Primary care pediatricians and pediatric subspecialists have a crucial role in the early consultation of a pediatric oncology center experienced in molecular diagnostics to facilitate a comprehensive evaluation for children with inflammatory tumors.

Single-Cell RNA-Seq Reveals Cellular Hierarchies and Impaired Developmental Trajectories in Pediatric Ependymoma.

Ependymoma is a heterogeneous entity of central nervous system tumors with well-established molecular groups. Here, we apply single-cell RNA sequencing to analyze ependymomas across molecular groups and anatomic locations to investigate their intratumoral heterogeneity and developmental origins. Ependymomas are composed of a cellular hierarchy initiating from undifferentiated populations, which undergo impaired differentiation toward three lineages of neuronal-glial fate specification. While prognostically favorable groups of ependymoma predominantly harbor differentiated cells, aggressive groups are enriched for undifferentiated cell populations. The delineated transcriptomic signatures correlate with patient survival and define molecular dependencies for targeted treatment approaches. Taken together, our analyses reveal a developmental hierarchy underlying ependymomas relevant to biological and clinical behavior.