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KIAA1549-BRAF Expression Establishes a Permissive Tumor Microenvironment Through NF?B-Mediated CCL2 Production.

ABSTRACT: KIAA1549-BRAF is the most frequently identified genetic mutation in sporadic pilocytic astrocytoma (PA), creating a fusion BRAF (f-BRAF) protein with increased BRAF activity. Fusion-BRAF-expressing neural stem cells (NSCs) exhibit increased cell growth and can generate glioma-like lesions following injection into the cerebella of naïve mice. Increased Iba1+ monocyte (microglia) infiltration is associated with murine f-BRAF-expressing NSC-induced glioma-like lesion formation, suggesting that f-BRAF-expressing NSCs attract microglia to establish a microenvironment supportive of tumorigenesis. Herein, we identify Ccl2 as the chemokine produced by f-BRAF-expressing NSCs, which is critical for creating a permissive stroma for gliomagenesis. In addition, f-BRAF regulation of Ccl2 production operates in an ERK- and NF?B-dependent manner in cerebellar NSCs. Finally, Ccr2-mediated microglia recruitment is required for glioma-like lesion formation in vivo, as tumor do not form in Ccr2-deficient mice following f-BRAF-expressing NSC injection. Collectively, these results demonstrate that f-BRAF expression creates a supportive tumor microenvironment through NF?B-mediated Ccl2 production and microglia recruitment.

SUBMITTER: Chen R

PROVIDER: S-EPMC6277251 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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KIAA1549-BRAF Expression Establishes a Permissive Tumor Microenvironment Through NFκB-Mediated CCL2 Production.

Chen Ran R Keoni Chanel C Waker Christopher A CA Lober Robert M RM Chen Yi-Hsien YH Gutmann David H DH

Neoplasia (New York, N.Y.) 20181130 1

KIAA1549-BRAF is the most frequently identified genetic mutation in sporadic pilocytic astrocytoma (PA), creating a fusion BRAF (f-BRAF) protein with increased BRAF activity. Fusion-BRAF-expressing neural stem cells (NSCs) exhibit increased cell growth and can generate glioma-like lesions following injection into the cerebella of naïve mice. Increased Iba1<sup>+</sup> monocyte (microglia) infiltration is associated with murine f-BRAF-expressing NSC-induced glioma-like lesion formation, suggestin ...[more]

PMID: 30504064

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Project description:Hypoxia is caused by a cancer-promoting milieu characterized by persistent inflammation. NF-κB and HIF-1α are critical participants in this transition. Tumor development and maintenance are aided by NF-κB, while cellular proliferation and adaptability to angiogenic signals are aided by HIF-1α. Prolyl hydroxylase-2 (PHD-2) has been hypothesized to be the key oxygen-dependent regulator of HIF-1α and NF-transcriptional B's activity. Without low oxygen levels, HIF-1α is degraded by the proteasome in a process dependent on oxygen and 2-oxoglutarate. As opposed to the normal NF-κB activation route, where NF-κB is deactivated by PHD-2-mediated hydroxylation of IKK, this method actually activates NF-κB. HIF-1α is protected from degradation by proteasomes in hypoxic cells, where it then activates transcription factors involved in cellular metastasis and angiogenesis. The Pasteur phenomenon causes lactate to build up inside the hypoxic cells. As part of a process known as lactate shuttle, MCT-1 and MCT-4 cells help deliver lactate from the blood to neighboring, non-hypoxic tumour cells. Non-hypoxic tumour cells use lactate, which is converted to pyruvate, as fuel for oxidative phosphorylation. OXOPHOS cancer cells are characterized by a metabolic switch from glucose-facilitated oxidative phosphorylation to lactate-facilitated oxidative phosphorylation. Although PHD-2 was found in OXOPHOS cells. There is no clear explanation for the presence of NF-kappa B activity. The accumulation of the competitive inhibitor of 2-oxo-glutarate, pyruvate, in non-hypoxic tumour cells is well established. So, we conclude that PHD-2 is inactive in non-hypoxic tumour cells due to pyruvate-mediated competitive suppression of 2-oxo-glutarate. This results in canonical activation of NF-κB. In non-hypoxic tumour cells, 2-oxoglutarate serves as a limiting factor, rendering PHD-2 inactive. However, FIH prevents HIF-1α from engaging in its transcriptional actions. Using the existing scientific literature, we conclude in this study that NF-κB is the major regulator of tumour cell growth and proliferation via pyruvate-mediated competitive inhibition of PHD-2.

Project description:Microvascular proliferation (MVP), an aberrant vascular structure containing multilayered mitotically active endothelial- and smooth-muscle cells/pericytes, is a histopathological hallmark of glioblastoma multiforme (GBM). Although MVP tends to be associated with high-grade glioma, it has also been detected in WHO grade I pilocytic astrocytoma (PA). However, little is known about the mechanism underlying its formation. Using TP53 point mutations as a marker for tumor-derived cells, we earlier reported that MVP was partially converted from tumor cells via mesenchymal transition. In the current study we used the KIAA1549-BRAF fusion gene as a marker to assess whether MVPs in PA contained tumor-derived cells and/or phenotypically distinct tumor cells expressing vascular markers. cDNA synthesized from frozen tissue of six PA patients operated at our institute was analyzed to detect the KIAA1549-BRAF fusion gene by reverse transcription polymerase chain reaction (RT-PCR) assay. The breakpoint in the fusion gene was identified by long and accurate PCR (LA-PCR) and Sanger sequencing of genomic DNA. Distinct tumor cells and cellular components of MVP were obtained by laser microdissection. For the qualitative and quantitative detection of the KIAA1549-BRAF fusion gene we performed genomic and digital PCR assays. Fluorescence in situ hybridization (FISH) was used to assess gene fusion in cellular components of MVP. Samples from three PA patients harbored the KIAA1549 exon 15, BRAF exon 9 fusion gene. In two patient samples with abundant MVP, RT-PCR assay detected strong bands arising from the KIAA1549-BRAF fusion gene in both tumor cells and cellular components of MVP. Digital PCR showed that vis-à-vis tumor tissue, its relative expression in cellular components of MVP was 42% in one- and 76% in another sample. FISH revealed amplified signals in both tumor cells and cellular components of MVP indicative of tandem duplication. Our findings suggest that in patients with PA, some cellular components of MVP contained tumor derived cell and/or phenotypically distinct tumor cells expressing vascular markers.

Dataset Information

KIAA1549-BRAF Expression Establishes a Permissive Tumor Microenvironment Through NF?B-Mediated CCL2 Production.

Publications

KIAA1549-BRAF Expression Establishes a Permissive Tumor Microenvironment Through NFκB-Mediated CCL2 Production.

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