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Only a subset of recurrent glioblastoma (rGBM) responds to anti-PD-1 immunotherapy. Previously, we reported enrichment of BRAF/PTPN11 mutations in 30% of rGBM that responded to PD-1 blockade. Given that BRAF and PTPN11 promote MAPK/ERK signaling, we investigated whether activation of this pathway is associated with response to PD-1 inhibitors in rGBM, including patients that do not harbor BRAF/PTPN11 mutations. Here we show that immunohistochemistry for ERK1/2 phosphorylation (p-ERK), a marker of MAPK/ERK pathway activation, is predictive of overall survival following adjuvant PD-1 blockade in two independent rGBM patient cohorts. Single-cell RNA-sequencing and multiplex immunofluorescence analyses revealed that p-ERK was mainly localized in tumor cells and that high-p-ERK GBMs contained tumor-infiltrating myeloid cells and microglia with elevated expression of MHC class II and associated genes. These findings indicate that ERK1/2 activation in rGBM is predictive of response to PD-1 blockade and is associated with a distinct myeloid cell phenotype.

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2009.01.026 Byline: Greg Holmes (a), Gerson Rothschild (a), Upal Basu Roy (a), Chu-Xia Deng (a)(b), Alka Mansukhani (a), Claudio Basilico (a) Keywords: Fgfr2; Craniosynostosis; Apert syndrome; Osteoblast; Coronal suture; Calvaria; Mouse; Intramembranous ossification Abstract: Activating mutations of FGFRs1-3 cause craniosynostosis (CS), the premature fusion of cranial bones, in man and mouse. The mechanisms by which such mutations lead to CS have been variously ascribed to increased osteoblast proliferation, differentiation, and apoptosis, but it is not always clear how these disturbances relate to the process of suture fusion. We have reassessed coronal suture fusion in an Apert Fgfr2 (S252W) mouse model. We find that the critical event of CS is the early loss of basal sutural mesenchyme as the osteogenic fronts, expressing activated Fgfr2, unite to form a contiguous skeletogenic membrane. A mild increase in osteoprogenitor proliferation precedes but does not accompany this event, and apoptosis is insignificant. On the other hand, the more apical coronal suture initially forms appropriately but then undergoes fusion, albeit at a slower rate, accompanied by a significant decrease in osteoprogenitor proliferation, and increased osteoblast maturation. Apoptosis now accompanies fusion, but is restricted to bone fronts in contact with one another. We correlated these in vivo observations with the intrinsic effects of the activated Fgfr2 S252W mutation in primary osteoblasts in culture, which show an increased capacity for both proliferation and differentiation. Our studies suggest that the major determinant of Fgfr2-induced craniosynostosis is the failure to respond to signals that would halt the recruitment or the advancement of osteoprogenitor cells at the sites where sutures should normally form. Author Affiliation: (a) Department of Microbiology, New York University School of Medicine, New York, NY 10016, USA (b) Genetics of Development and Disease Branch, National Institute of Diabetes, Digestive and Kidney Diseases, US National Institutes of Health, 10 Center Drive, Bethesda, Maryland 20892, USA Article History: Received 13 November 2008; Revised 16 January 2009; Accepted 20 January 2009

Author(s): Evangelos Pefanis [1, 2, 4]; Jiguang Wang [1, 3, 4]; Gerson Rothschild [1, 4]; Junghyun Lim [1]; Jaime Chao [1]; Raul Rabadan (corresponding author) [3]; Aris N. Economides [2]; [...]

Integrin-β1-null keratinocytes can adhere to fibronectin through integrin αvβ6, but form large peripheral focal adhesions and exhibit defective cell spreading. Here we report that, in addition to the reduced avidity of αvβ6 integrin binding to fibronectin, the inability of integrin β6 to efficiently bind and recruit kindlin-2 to focal adhesions directly contributes to these phenotypes. Kindlins regulate integrins through direct interactions with the integrin-β cytoplasmic tail and keratinocytes express kindlin-1 and kindlin-2. Notably, although both kindlins localize to focal adhesions in wild-type cells, only kindlin-1 localizes to the integrin-β adhesions of integrin-β cells. Rescue of these cells with wild-type and chimeric integrin constructs revealed a correlation between kindlin-2 recruitment and cell spreading. Furthermore, despite the presence of kindlin-1, knockdown of kindlin-2 in wild-type keratinocytes impaired cell spreading. Our data reveal unexpected functional consequences of differences in the association of two homologous kindlin isoforms with two closely related integrins, and suggest that despite their similarities, different kindlins are likely to have unique functions. [ABSTRACT FROM AUTHOR]

The inhibitor of differentiation 1d2 is a target of the retinoblastoma (Rb) protein during mouse embryogenesis. In Rb+/- mice, LOH at the wild-type Rb allele initiates pituitary adenocarcinoma, a tumor derived from embryonic melanotropes. Here we identically a critical role for Id2 in initiation, growth, and angiogenesis of pituitary tumors from Rb+/- mice. We show that proliferation and differentiation are intimately coupled in Rb+/- pituitary cells before tumor initiation. In Id2-null pituitaries, premature activation of basic helix-loop- helix-mediated transcription and expression of the cdk inhibitor p27Kip1 impairs the proliferation of melanotropes and tumor initiation. Without Id2, Rb+/- mice have fewer early tumor lesions and a markedly decreased proliferation rate of the tumor foci. Expression of Id2 by pituitary tumor cells promotes growth and angiogenesis by functioning as a master regulator of vascular endothelial growth factor (VEGF). In human neuroblastoma, the N-Myc-driven expression of 1d2 is sufficient and necessary for expression of VEGF. These results establish that aberrant 1d2 activity directs initiation and progression of embryonal cancer. [ABSTRACT FROM AUTHOR]

The Kit receptor tyrosine kinase functions in hematopoiesis, melanogenesis and gametogenesis. Kit receptor-mediated cellular responses include proliferation, survival, adhesion, secretion and differentiation. In mast cells, Kit-mediated recruitment and activation of phosphatidylinositol 3′-kinase (PI 3-kinase) produces phosphatidylinositol 3′-phosphates, plays a critical role in mediating cell adhesion and secretion and has contributory roles in mediating cell survival and proliferation. To investigate the consequences in vivo of blocking Kit-mediated PI 3-kinase activation we have mutated the binding site for the p85 subunit of PI 3-kinase in the Kit gene, using a knock-in strategy. Mutant mice have no pigment deficiency or impairment of steady-state hematopoiesis. However, gametogenesis is affected in several ways and tissue mast cell numbers are affected differentially. While primordial germ cells during embryonic development are not affected, KitY719F/KitY719F males are sterile due to a block at the premeiotic stages in spermatogenesis. Furthermore, adult males develop Leydig cell hyperplasia. The Leydig cell hyperplasia implies a role for Kit in Leydig cell differentiation and/or steroido-genesis. In mutant females follicle development is impaired at the cuboidal stages resulting in reduced fertility. Also, adult mutant females develop ovarian cysts and ovarian tubular hyperplasia. Therefore, a block in Kit receptor-mediated PI 3-kinase signaling may be compensated for in hematopoiesis, melanogenesis and primordial germ cell development, but is critical in spermatogenesis and oogenesis. [ABSTRACT FROM AUTHOR]

A precise balance between proliferation and differentiation must be maintained during neural development to obtain the correct proportion of differentiated cell types in the adult nervous system. The basic helix-loop-helix (bHLH) transcription factors known as E proteins and their natural inhibitors, the Id proteins, control the timing of differentiation and terminal exit from the cell cycle. Here we show that progression into S phase of human neuroblastoma cells is prevented by E proteins and promoted by Id2. Cyclin-dependent kinase inhibitors (CKI) have been identified as key effectors of cell cycle arrest in differentiating cells. However, p57Kip2 is the only CKI that is absolutely required for normal development. Through the use of global gene expression analysis in neuroblastoma cells engineered to acutely express the E protein E47 and Id2, we find that p57Kip2 is a target of E47. Consistent with the role of Id proteins, Id2 prevents activation of p57Kip2 expression, and the retinoblastoma tumor suppressor protein, a known Id2 inhibitor, counters this activity. The strong E47-mediated inhibition of entry into S phase is entirely reversed in cells in which expression of p57Kip2 is silenced by RNA interference. During brain development, expression of p57Kip2 is opposite that of Id2. Our findings identify p57Kip2 as a functionally relevant target recruited by bHLH transcription factors to induce cell cycle arrest in developing neuroblasts and suggest that deregulated expression of Id proteins may be an epigenetic mechanism to silence expression of this CKI in neural tumors. [ABSTRACT FROM AUTHOR]

Immunoglobulin heavy chain (IgH) locus-associated G-rich long noncoding RNA (SμGLT) is important for physiological and pathological B cell DNA recombination. We demonstrate that the METTL3 enzyme-catalyzed N 6 -methyladenosine (m 6 A) RNA modification drives recognition and 3' end processing of SμGLT by the RNA exosome, promoting class switch recombination (CSR) and suppressing chromosomal translocations. The recognition is driven by interaction of the MPP6 adaptor protein with nuclear m 6 A reader YTHDC1. MPP6 and YTHDC1 promote CSR by recruiting AID and the RNA exosome to actively transcribe SμGLT. Direct suppression of m 6 A modification of SμGLT or of m 6 A reader YTHDC1 reduces CSR. Moreover, METTL3, an essential gene for B cell development in the bone marrow and germinal center, suppresses IgH-associated aberrant DNA breaks and prevents genomic instability. Taken together, we propose coordinated and central roles for MPP6, m 6 A modification, and m 6 A reader proteins in controlling long noncoding RNA processing, DNA recombination, and development in B cells., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Purpose: Cancer immunoediting shapes tumor progression by the selection of tumor cell variants that can evade immune recognition. Given the immune evasion and intratumor heterogeneity characteristic of gliomas, we hypothesized that CD8 + T cells mediate immunoediting in these tumors., Experimental Design: We developed retrovirus-induced PDGF + Pten -/- murine gliomas and evaluated glioma progression and tumor immunogenicity in the absence of CD8 + T cells by depleting this immune cell population. Furthermore, we characterized the genomic alterations present in gliomas that developed in the presence and absence of CD8 + T cells., Results: Upon transplantation, gliomas that developed in the absence of CD8 + T cells engrafted poorly in recipients with intact immunity but engrafted well in those with CD8 + T-cell depletion. In contrast, gliomas that developed under pressure from CD8 + T cells were able to fully engraft in both CD8 + T-cell-depleted mice and immunocompetent mice. Remarkably, gliomas developed in the absence of CD8 + T cells exhibited increased aneuploidy, MAPK pathway signaling, gene fusions, and macrophage/microglial infiltration, and showed a proinflammatory phenotype. MAPK activation correlated with macrophage/microglia recruitment in this model and in the human disease., Conclusions: Our studies indicate that, in these tumor models, CD8 + T cells influence glioma oncogenic pathways, tumor genotype, and immunogenicity. This suggests immunoediting of immunogenic tumor clones through their negative selection by CD8 + T cells during glioma formation., (©2020 American Association for Cancer Research.)