PDGF Signaling Research Articles

Long Non-Coding RNA Expression Profile Associated with Malignant Progression of Oral Submucous Fibrosis.

Oral submucous fibrosis (OSF) as one of the premalignant disorders endures a series of histopathological stages to invasive oral squamous cell carcinoma (OSCC) eventually. However, the role of long non-coding RNA (lncRNA) expression in OSF malignant progression still remains poorly understood. Through RNA-sequencing normal mucous, OSF and OSCC tissues, we found 687 lncRNA transcripts significantly and differentially expressed during OSF progression, including 231 upregulated lncRNAs and 456 downregulated lncRNAs, indicating that lncRNAs are involved in the regulation of different stages of OSF development. Further functional enrichment analysis showed these differentially expressed lncRNAs participated in inflammation signaling, Wnt signaling, angiogenesis, CCKR signaling, integrin signaling, PDGF signaling, p53 signaling, and EGF receptor (EGFR) signaling pathways, which contribute to inflammatory and fibroelastic pathogenetic changes of OSF and further malignant progression. Five novel lncRNAs were differentially expressed during OSF progression with varied expression levels, indicating the importance of these lncRNAs in OSF malignant development. Moreover, some lncRNAs have been previously identified to be associated with OSCC pathogenesis, including HCG22, RP11-397A16.1, LINC00271, CTD-3179P9.1, and ZNF667-AS1. Thus, our study firstly comprehensively elucidated lncRNAs expression profile of malignant procession from OSF premalignant lesion to OSCC, which will enlighten our understanding of the importance of lncRNA involved in OSF malignant development.

Angiocrine signals regulate quiescence and therapy resistance in bone metastasis.

Bone provides supportive microenvironments for hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs) and is a frequent site of metastasis. While incidences of bone metastases increase with age, the properties of the bone marrow microenvironment that regulate dormancy and reactivation of disseminated tumor cells (DTCs) remain poorly understood. Here, we elucidate the age-associated changes in the bone secretome that trigger proliferation of HSCs, MSCs, and DTCs in the aging bone marrow microenvironment. Remarkably, a bone-specific mechanism involving expansion of pericytes and induction of quiescence-promoting secretome rendered this proliferative microenvironment resistant to radiation and chemotherapy. This bone-specific expansion of pericytes was triggered by an increase in PDGF signaling via remodeling of specialized type H blood vessels in response to therapy. The decline in bone marrow pericytes upon aging provides an explanation for loss of quiescence and expansion of cancer cells in the aged bone marrow microenvironment. Manipulation of blood flow — specifically, reduced blood flow — inhibited pericyte expansion, regulated endothelial PDGF-B expression, and rendered bone metastatic cancer cells susceptible to radiation and chemotherapy. Thus, our study provides a framework to recognize bone marrow vascular niches in age-associated increases in metastasis and to target angiocrine signals in therapeutic strategies to manage bone metastasis.

An oncogenic activity of PDGF-C and its splice variant in human breast cancer

Despite strong evidence for the involvement of PDGF signaling in breast cancer, little is known about the PDGF ligand responsible for PDGFR activation during breast cancer progression. Here, we found PDGF-C to be highly expressed in breast carcinoma cell lines. Immunohistochemical analysis of invasive breast cancer revealed an association between increased PDGF-C expression and lymph node metastases, Ki-67 proliferation index, and poor disease-free survival. We also identified a PDGF-C splice variant encoding truncated PDGF-C (t-PDGF-C) isoform lacking the signal peptide and the N-terminal CUB domain. While t-PDGF C homodimer is retained intracellularly, it can be secreted as a heterodimer with full-length PDGF-C (FL-PDGF-C). PDGF-C downregulation reduced anchorage-independent growth and matrigel invasion of MDA-MB-231 cells. Conversely, ectopic expression of t-PDGF-C enhanced phenotypic transformation and invasion in BT-549 cells expressing endogenous FL-PDGF-C. The present study provides new insights into the functional significance of PDGF-C and its splice variant in human breast cancer.

Abstract 860: Solution structure of a dGMP fill-in G-quadruplex forms in PDGFR-b promoter

Abstract Platelet-derived growth factor receptor beta (PDGFR-β) is an important cell-surface-receptor tyrosine kinase implicated in PDGF signaling pathways. The PDGFR-β signaling pathway has been demonstrated as an essential component in control of the cellular growth, proliferation, survival, motility and differentiation. Overexpression of PDGFR-β is associated with tumor growth, angiogenesis, and migration, making PDGFR-β an attractive target for anticancer therapeutics. In the PDGFR-β gene promoter, a highly GC-rich proximal promoter region -165 to -139 nt upstream of the transcriptional start site is crucial for basal promoter activity, which can form G-quadruplexes (G4s). These structures can be stabilized by the G4-interactive molecules to inhibit the PDGFR-β transcriptional activity. Our previous structure study showed that the G4 structures formed in the PDGFR-β gene promoter always contain a broken G-strand. The broken-strand structure suggests a possibility of the formation of a G-vacancy-bearing G-quadruplex (GVBQ) structure, which can be complemented and stabilized by guanine and guanine derivatives. Elevated levels of guanine and guanine derivatives/metabolites are found in tumor cells, suggesting that the GVBQs may play important regulatory roles in tumor cell pathological processes, such as PDGFR-β transcriptional regulation. Using NMR, CD, EMSA and DMS footprinting experiments, herein, we contributed the first atomic resolution structure of a dGMP fill-in G4 formed in PDGFR-β gene promoter in potassium solution. Surprisingly, our results showed that solution dGMP preferentially inserts from the 5' end tetrad instead of the G-22 position of the previously determined structure, and maybe in dynamic equilibrium with our previously reported G-quadruplex. The dGMP, like other intramolecular guanines, that forms four Hoogsteen hydrogen bonds with adjacent guanines and stabilizes by electronic coordination with potassium cation as well as the π-π stacking interaction with the middle tetrad guanine. Moreover, a series of guanine derivatives were investigated to fill in this GVBQ, and found that only these derivatives that keep the ability to form Hoogsteen hydrogen bonds in G-tetrad can serve as the substrate, providing insights into designing new guanine derivatives anticancer drugs. Citation Format: Kaibo Wang, Jonathan Dickerhoff, Guanhui Wu, Clement Lin, Danzhou Yang. Solution structure of a dGMP fill-in G-quadruplex forms in PDGFR-b promoter [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 860.

Fibrosis driving myofibroblast precursors in MPN and new therapeutic pathways.

Fibrosis driving myofibroblast precursors in MPN and new therapeutic pathways.

Growth Factor Signaling Regulates Mechanical Nociception in Flies and Vertebrates.

Mechanical sensitization is one of the most difficult clinical pain problems to treat. However, the molecular and genetic bases of mechanical nociception are unclear. Here we develop a Drosophila model of mechanical nociception to investigate the ion channels and signaling pathways that regulate mechanical nociception. We fabricated von Frey filaments that span the subthreshold to high noxious range for Drosophila larvae. Using these, we discovered that pressure (force/area), rather than force per se, is the main determinant of aversive rolling responses to noxious mechanical stimuli. We demonstrated that the RTK PDGF/VEGF receptor (Pvr) and its ligands (Pvfs 2 and 3) are required for mechanical nociception and normal dendritic branching. Pvr is expressed and functions in class IV sensory neurons, whereas Pvf2 and Pvf3 are produced by multiple tissues. Constitutive overexpression of Pvr and its ligands or inducible overexpression of Pvr led to mechanical hypersensitivity that could be partially separated from morphological effects. Genetic analyses revealed that the Piezo and Pain ion channels are required for mechanical hypersensitivity observed upon ectopic activation of Pvr signaling. PDGF, but not VEGF, peptides caused mechanical hypersensitivity in rats. Pharmacological inhibition of VEGF receptor Type 2 (VEGFR-2) signaling attenuated mechanical nociception in rats, suggesting a conserved role for PDGF and VEGFR-2 signaling in regulating mechanical nociception. VEGFR-2 inhibition also attenuated morphine analgesic tolerance in rats. Our results reveal that a conserved RTK signaling pathway regulates baseline mechanical nociception in flies and rats.SIGNIFICANCE STATEMENT Hypersensitivity to touch is poorly understood and extremely difficult to treat. Using a refined Drosophila model of mechanical nociception, we discovered a conserved VEGF-related receptor tyrosine kinase signaling pathway that regulates mechanical nociception in flies. Importantly, pharmacological inhibition of VEGF receptor Type 2 signaling in rats causes analgesia and blocks opioid tolerance. We have thus established a robust, genetically tractable system for the rapid identification and functional analysis of conserved genes underlying mechanical pain sensitivity.

PDGF-B: The missing piece in the mosaic of PDGF family role in craniofacial development.

The platelet-derived growth factor (PDGF) family consists of four ligands (PDGF-A, PDGF-B, PDGF-C, PDGF-D) and two tyrosine kinase receptors (PDGFR-α and PDGFR-β). In vertebrates, PDGF signaling influences cell proliferation, migration, and matrix deposition, and its up-regulation is implicated in cancer progression. Despite this evidence, the role of each family member during embryogenesis is still incomplete and partially controversial. In particular, study of the role of pdgf signaling during craniofacial development has been focused on pdgf-a, while the role of pdgf-b is almost unknown due to the lethal phenotypes of pdgf-b-null mice. By using a pdgf-b splice-blocking morpholino approach, we highlighted impairment of neural crest cell (NCC) migration in Xenopus laevis morphants, leading to alteration of NCC derivatives formation, such as cranial nerves and cartilages. We also uncovered a possible link between pdgf-b and the expression of cadherin superfamily members cdh6 and cdh11, which mediate cell-cell adhesion promoting NCC migration. Our results suggested that pdgf-b signaling is involved in cranial NCC migration and it is required for proper formation of craniofacial NCC derivatives. Taken together, these data unveiled a new role for pdgf-b during vertebrate development, contributing to complete the picture of pdgf signaling role in craniofacial development.

Heparin Differentially Impacts Gene Expression of Stromal Cells from Various Tissues

Pooled human platelet lysate (pHPL) is increasingly used as replacement of animal serum for manufacturing of stromal cell therapeutics. Porcine heparin is commonly applied to avoid clotting of pHPL-supplemented medium but the influence of heparin on cell behavior is still unclear. Aim of this study was to investigate cellular uptake of heparin by fluoresceinamine-labeling and its impact on expression of genes, proteins and function of human stromal cells derived from bone marrow (BM), umbilical cord (UC) and white adipose tissue (WAT). Cells were isolated and propagated using various pHPL-supplemented media with or without heparin. Flow cytometry and immunocytochemistry showed differential cellular internalization and lysosomal accumulation of heparin. Transcriptome profiling revealed regulation of distinct gene sets by heparin including signaling cascades involved in proliferation, cell adhesion, apoptosis, inflammation and angiogenesis, depending on stromal cell origin. The influence of heparin on the WNT, PDGF, NOTCH and TGFbeta signaling pathways was further analyzed by a bead-based western blot revealing most alterations in BM-derived stromal cells. Despite these observations heparin had no substantial effect on long-term proliferation and in vitro tri-lineage differentiation of stromal cells, indicating compatibility for clinically applied cell products.

PDGF Signaling in Primitive Endoderm Cell Survival Is Mediated by PI3K-mTOR Through p53-Independent Mechanism.

Receptor tyrosine kinase signaling pathways are key regulators for the formation of the primitive endoderm (PrE) and the epiblast (Epi) from the inner cell mass (ICM) of the mouse preimplantation embryo. Among them, FGF signaling is critical for PrE cell specification, whereas PDGF signaling is critical for the survival of committed PrE cells. Here, we investigated possible functional redundancies among FGF, PDGF, and KIT signaling and showed that only PDGF signaling is involved in PrE cell survival. In addition, we analyzed the effectors downstream of PDGFRα. Our results suggest that the role of PDGF signaling in PrE cell survival is mediated through PI3K-mTOR and independently from p53. Lastly, we uncovered a role for PI3K-mTOR signaling in the survival of Epi cells. Taken together, we propose that survival of ICM cell lineages relies on the regulation of PI3K-mTOR signaling through the regulation of multiple signaling pathways. Stem Cells 2019;37:888-898.

Integrating germline and somatic variation information using genomic data for the discovery of biomarkers in prostate cancer

BackgroundProstate cancer (PCa) is the most common diagnosed malignancy and the second leading cause of cancer-related deaths among men in the United States. High-throughput genotyping has enabled discovery of germline genetic susceptibility variants (herein referred to as germline mutations) associated with an increased risk of developing PCa. However, germline mutation information has not been leveraged and integrated with information on acquired somatic mutations to link genetic susceptibility to tumorigenesis. The objective of this exploratory study was to address this knowledge gap.MethodsGermline mutations and associated gene information were derived from genome-wide association studies (GWAS) reports. Somatic mutation and gene expression data were derived from 495 tumors and 52 normal control samples obtained from The Cancer Genome Atlas (TCGA). We integrated germline and somatic mutation information using gene expression data. We performed enrichment analysis to discover molecular networks and biological pathways enriched for germline and somatic mutations.ResultsWe discovered a signature of 124 genes containing both germline and somatic mutations. Enrichment analysis revealed molecular networks and biological pathways enriched for germline and somatic mutations, including, the PDGF, P53, MYC, IGF-1, PTEN and Androgen receptor signaling pathways.ConclusionIntegrative genomic analysis links genetic susceptibility to tumorigenesis in PCa and establishes putative functional bridges between the germline and somatic variation, and the biological pathways they control.

Lesion Dynamics Under Varying Paracrine PDGF Signaling in Brain Tissue.

Paracrine PDGF signaling is involved in many processes in the body, both normal and pathological, including embryonic development, angiogenesis, and wound healing as well as liver fibrosis, atherosclerosis, and cancers. We explored this seemingly dual (normal and pathological) role of PDGF mathematically by modeling the release of PDGF in brain tissue and then varying the dynamics of this release. Resulting simulations show that by varying the dynamics of a PDGF source, our model predicts three possible outcomes for PDGF-driven cellular recruitment and lesion growth: (1) localized, short duration of growth, (2) localized, chronic growth, and (3) widespread chronic growth. Further, our model predicts that the type of response is much more sensitive to the duration of PDGF exposure than the maximum level of that exposure. This suggests that extended duration of paracrine PDGF signal during otherwise normal processes could potentially lead to lesions having a phenotype consistent with pathologic conditions.

Combined Notch and PDGF Signaling Enhances Migration and Expression of Stem Cell Markers while Inducing Perivascular Cell Features in Muscle Satellite Cells.

SummarySatellite cells are responsible for skeletal muscle regeneration. Upon activation, they proliferate as transient amplifying myoblasts, most of which fuse into regenerating myofibers. Despite their remarkable differentiation potential, these cells have limited migration capacity, which curtails clinical use for widespread forms of muscular dystrophy. Conversely, skeletal muscle perivascular cells have less myogenic potential but better migration capacity than satellite cells. Here we show that modulation of Notch and PDGF pathways, involved in developmental specification of pericytes, induces perivascular cell features in adult mouse and human satellite cell-derived myoblasts. DLL4 and PDGF-BB-treated cells express markers of perivascular cells and associate with endothelial networks while also upregulating markers of satellite cell self-renewal. Moreover, treated cells acquire trans-endothelial migration ability while remaining capable of engrafting skeletal muscle upon intramuscular transplantation. These results extend our understanding of muscle stem cell fate plasticity and provide a druggable pathway with clinical relevance for muscle cell therapy.

Paracrine Mechanisms Involved in Mesenchymal Stem Cell Differentiation into Cardiomyocytes.

Cardiovascular Disease (CVD) is one of the world-wide healthcare problem that involves the heart or blood vessels. CVD includes myocardial infarction and coronary artery diseases (CAD). Dysfunctional myocardial cells are leading causes of low cardiac output or ventricular dysfunction after cardiac arrest and may contribute to the progression of CVD which could not generate new cardiomyocytes in human adult heart. The mesenchymal stem cells (MSCs) which are present in adult marrow can self-renew and have the capacity of differentiation into multiple types of cells including cardiomyocytes. Recent biochemical analyses greatly revealed that several regulators of MSCs, such as HGF, PDGF, Wnt, and Notch-1 signaling pathways have been shown to be involved in the proliferation and differentiation into cardiomyocytes. Preclinical studies are paving the way for further applications of MSCs in the repair of myocardial infarction. In this study, we discuss and summarize the paracrine mechanisms involved in MSCs differentiation into cardiomyocytes.

CD166 Engagement Augments Mouse and Human Hematopoietic Stem and Progenitor Function Via Activation of Stem Cell-Associated and Cell Cycle Pathways

Functions of hematopoietic stem and progenitor cells are regulated by cellular signaling networks and by the cellular and non-cellular elements of the hematopoietic niche. CD166 is highly expressed on both human and murine hematopoietic stem cells (HSC) and on murine osteoblasts and has been identified as a critical regulator of their functions. CD166 engages in trans-homophilic interactions with other CD166 molecules. However how homophilic or heterophilic engagement of CD166 (with CD6) improves HSC function is unknown.Since we described that CD166 is expressed on murine and human HSC and murine osteoblasts, we hypothesized that CD166-CD166 homophilic engagement is critical for the hematopoiesis enhancing activity (HEA) that is mediated by osteoblasts. CD166+LSK cells cultured for 7d on either WT osteoblasts or recombinant murine CD166 (rmCD166) showed increased colony forming units (CFU) compared to CD166-LSK cells cultured identically (p<0.05). Moreover, after 7d, the frequency of Lin-Sca1+ cells was higher in co-cultures containing rmCD166 and CD166+LSK than cultures initiated with CD166-LSK (p<0.05).Next, we determined the effect of loss of CD166 expression on progenitors and osteoblasts on hematopoietic functions using CD166 knockout mice (CD166-/-). CD166-/- LSK cells co-cultured with rmCD166 showed decreased CFU production compared to WT LSK cultured similarly (p<0.05). In addition, WT HSC cultured on CD166-/- osteoblasts for 7d showed decreased fold change in CFU relative to WT HSC cultured over WT osteoblasts (p<0.05). In vivo transplantation studies to corroborate our in vitro results are ongoing. To determine if CD166-CD166 interactions also enhance hematopoietic functions of human HSC, we cultured CD34+CD166+ and CD34+CD166- HSC isolated from cord blood with recombinant human CD166 (rhCD166). On d7, CD34+CD166+ HSC cultured with rhCD166 showed 30±4-fold increase in CFU production compared to d0, whereas CD34+CD166- HSC had only 9±0.4-fold increase. Taken together, these data suggest that CD166-CD166 homophilic interactions enhance hematopoietic functions and loss of this homophilic interaction negatively impacts HSC.To elucidate the underlying signaling mechanism of CD166-CD166 mediated HEA, we cultured WT or CD166-/- SLAM LSK cells with rmCD166 for 20hr and performed single-cell (sc) RNA seq. We analyzed for differential gene expression (DEG) using LGMT model and identified 518 upregulated and 174 downregulated genes in CD166-/- HSC compared to WT. Following pathway enrichment analysis of DEGs, we identified 148 canonical pathways enriched by the upregulated genes in CD166-/- HSC, including cell cycle, translational regulation, and mitochondria-related signaling pathways. 268 pathways were impacted by the downregulated genes in CD166-/- HSC, including oxidative stress response, and metabolism. Moreover, CXCR4 signaling, PDGF signaling and glucocorticoid receptor signaling pathways were also downregulated in CD166-/- HSC.A cell trajectory reflecting associations among cells revealed a single cluster of CD166-/- HSC. CD166-/- HSC are linked with low expression of stemness marker genes, and high expression of genes regulating cell cycle, oxidative phosphorylation, and glucose metabolism. In addition, ER-stress and oxidative stress responsive genes are overexpressed in CD166-/- HSC. In CD166-/- HSC, genes of all enriched pathways were highly connected in the co-expressed networks, which indicates that in HSC, the impact of loss of CD166 on cell cycle, metabolism, growth factors and stemness pathways is highly associated. Sixteen hub genes including Suclg1, Eif4a1, Cox4i2, Jak3, Runx3, and Cdk6 were identified in the co-expression network.We next applied bi-clustering algorithm QUBIC to identify modules of co-upregulated genes to analyze the transcriptomic variations of DEG in CD166-/- HSC and identified 39-gene co-upregulation modules forming one big block in CD166-/- HSC and 35-gene modules forming three blocks in WT HSC indicating that modules closely corresponded to CD166's impact over different cell states rather than on individual pathways.Overall, our studies suggest that homophilic CD166 interactions involving HSC are required for maintenance of essential pathways that sustain HSC function and progenitor cell production including stemness, mitochondrial function, metabolism, cell cycle and growth factor signaling. DisclosuresNo relevant conflicts of interest to declare.

Gene Expression Signatures Point to a Male Sex-Specific Lung Mesenchymal Cell PDGF Receptor Signaling Defect in Infants Developing Bronchopulmonary Dysplasia

Male sex is a risk factor for development of bronchopulmonary dysplasia (BPD), a common chronic lung disease following preterm birth. We previously found that tracheal aspirate mesenchymal stromal cells (MSCs) from premature infants developing BPD show reduced expression of PDGFRα, which is required for normal lung development. We hypothesized that MSCs from male infants developing BPD exhibit a pathologic gene expression profile deficient in PDGFR and its downstream effectors, thereby favoring delayed lung development. In a discovery cohort of 6 male and 7 female premature infants, we analyzed the tracheal aspirate MSCs transcriptome. A unique gene signature distinguished MSCs from male infants developing BPD from all other MSCs. Genes involved in lung development, PDGF signaling and extracellular matrix remodeling were differentially expressed. We sought to confirm these findings in a second cohort of 13 male and 12 female premature infants. mRNA expression of PDGFRA, FGF7, WNT2, SPRY1, MMP3 and FOXF2 were significantly lower in MSCs from male infants developing BPD. In female infants developing BPD, tracheal aspirate levels of proinflammatory CCL2 and profibrotic Galectin-1 were higher compared to male infants developing BPD and female not developing BPD. Our findings support a notion for sex-specific differences in the mechanisms of BPD development.

Impact of vitamin D depletion during development on mouse sperm DNA methylation

ABSTRACTSuboptimal environmental conditions during development can substantially alter the epigenome. Stable environmentally-induced changes to the germline epigenome, in particular, have important implications for the health of the next generation. We showed previously that developmental vitamin D depletion (DVD) resulted in loss of DNA methylation at several imprinted loci over two generations. Here, we assessed the impact of DVD on genome-wide methylation in mouse sperm in order to characterize the number, extent and distribution of methylation changes in response to DVD and to find genes that may be susceptible to this prevalent environmental perturbation. We detected 15,827 loci that were differentially methylated in DVD mouse sperm vs. controls. Most epimutations (69%) were loss of methylation, and the extent of methylation change and number of CpGs affected in a region were associated with genic location and baseline methylation state. Methylation response to DVD at validated loci was only detected in offspring that exhibited a phenotypic response to DVD (increased body and testes weight) suggesting the two types of responses are linked, though a causal relationship is unclear. Epimutations localized to regions enriched for developmental and metabolic genes and pathway analyses showed enrichment for Cadherin, Wnt, PDGF and Integrin signaling pathways. These findings show for the first time that vitamin D status during development leads to substantial DNA methylation changes across the sperm genome and that locus susceptibility is linked to genomic and epigenomic context.

Distinct mechanisms for PDGF and FGF signaling in primitive endoderm development

FGF signaling is known to play a critical role in the specification of primitive endoderm (PrE) and epiblast (Epi) from the inner cell mass (ICM) during mouse preimplantation development, but how FGFs synergize with other growth factor signaling pathways is unknown. Because PDGFRα signaling has also been implicated in the PrE, we investigated the coordinate functions of PDGFRα together with FGFR1 or FGFR2 in PrE development. PrE development was abrogated in Pdgfra; Fgfr1 compound mutants, or significantly reduced in Pdgfra; Fgfr2 or PdgfraPI3K; Fgfr2 compound mutants. We provide evidence that both Fgfr2 and Pdgfra play roles in PrE cell survival while Fgfr1 controls PrE cell specification. Our results suggest a model where FGFR1-engaged ERK1/2 signaling governs PrE specification while PDGFRα- and by analogy possibly FGFR2- engaged PI3K signaling regulates PrE survival and positioning in the embryo. Together, these studies indicate how multiple growth factors and signaling pathways can cooperate in preimplantation development.

Abstract 2480: Olig2-dependent reciprocal shift in PDGF and EGF receptor signaling regulates tumor phenotype and mitotic growth in malignant glioma

Abstract Malignant gliomas exhibit extensive heterogeneity and poor prognosis. Diverse molecular characteristics in gliomas represent a significant hurdle for effective therapy. We uncover that a population of Olig2+ cells function as glioma-propagating cells and is critical for tumor initiation of primary gliomas in mice. Elimination of Olig2+ mitotic progenitors blocks tumor initiation and progression. Intriguingly, deletion of Olig2 resulted in tumors that grow, albeit at a decelerated rate. Genome occupancy and expression profiling analyses reveal that OLIG2 directly activates cell proliferation machinery to promote tumorigenesis. Olig2 deletion causes a tumor phenotypic shift from an oligodendrocyte precursor-correlated proneural toward an astrocyte-associated gene expression pattern, manifest in downregulation of PDGF receptor-alpha and reciprocal upregulation of EGFR. Olig2 deletion further sensitizes glioma cells to EGFR inhibitors and extends animal lifespans. Our observations reveal that OLIG2 is a molecular arbiter of tumor phenotype plasticity that may underlie drug resistance, suggesting new strategies for enhancing EGFR drug sensitivity in glioma treatment. Citation Format: Fanghui Lu, Mei Xin, Qing Richard Lu. Olig2-dependent reciprocal shift in PDGF and EGF receptor signaling regulates tumor phenotype and mitotic growth in malignant glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2480.

Abstract 49: Stromal platelet derived growth factor receptor (PDGFRβ) signaling: A novel therapeutic target for breast cancer brain metastasis (BCBM)

Abstract Stromal PDGFRβ has emerged as an actionable mediator of breast tumor-stromal communication. As a receptor tyrosine kinase, PDGFRβ is activated by its ligand, PDGFB, which is released by neighboring tumor epithelium and endothelium. However, how PDGF signaling mediates breast cancer initiation, progression, and metastasis remains unclear. To evaluate PDGFRβ in this disease, we developed a mouse model of stromal-specific PDGFRβ activation using the Fsp-cre transgene previously published by our group (PDGFRβ mutant). PDGFRβ mutant mammary glands exhibit increased tertiary side-branching and epithelial proliferation confirming a stromal-specific PDGFRβ effect on neighboring epithelium. To evaluate the functional relevance of PDGFRβ activation on metastatic progression, we performed tail vein injection of PDGFB expressing murine mammary tumor cells and, surprisingly, observed brain metastases in 50% of the PDGFRβ mutant mice while no brain lesions were seen in controls. There was no difference in the incidence of lung, liver or bone metastases. Mammary tumor cells expressing low PDGFB did not exhibit a similar increase in brain metastases in mutant mice. While there is no observable difference in blood brain barrier permeability in the mutant mice, we bypassed this variable by intracranially injecting mammary tumor cells and found that larger tumors formed in the brains of PDGFRβ mutant mice versus controls. To our knowledge, this is the first example where genetic manipulation of the stroma leads to an increased incidence of BCBM. Also, our pre-clinical data suggests that primary breast tumors that express high PDGFB could preferentially metastasize to the brain. To test this in patients, we analyzed PDGFB protein expression in a tissue microarray comprised of HER2-positive and triple negative breast cancer (TNBC) primary tumors. While high PDGFB did not correlate with site-independent metastatic recurrence, it was prognostic of brain metastasis, mirroring our mouse data. Evaluation of PDGFB in a small cohort of matched primary breast tumors with associated brain (n=5) and lung metastases (n=2) revealed intense PDGFB staining in 100% of the brain metastases, but only 50% of the lung metastases. Our findings suggest high primary tumor PDGFB expression defines a subset of breast cancer patients predisposed to brain metastases. These patients may benefit from therapeutic intervention of PDGFRβ signaling. To test this pre-clinically, we treated mice harboring intracranial tumors with the PDGFR specific inhibitor, Crenolanib. Excitingly, Crenolanib treatment significantly inhibited the brain tumor burden in these mice. Combined, our findings (1) advocate that primary tumor expression of PDGFB is a novel prognostic biomarker for the development of BCBM and (2) support clinical trial evaluation of PDGFR inhibitors for the prevention and treatment of BCBM. Citation Format: Katie A. Thies, Anisha M. Hammer, B. Eason Hildreth, Luke O. Russell, Steven T. Sizemore, Anthony J. Trimboli, Raleigh D. Kladney, Chelsea M. Bolyard, Robert Pilarski, Lynn Schoenfield, Jose Otero, Arnab Chakravarti, Matthew Ringel, Balveen Kaur, Gustavo Leone, Michael C. Ostrowski, Gina M. Sizemore. Stromal platelet derived growth factor receptor (PDGFRβ) signaling: A novel therapeutic target for breast cancer brain metastasis (BCBM) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 49.

Trapidil induces osteogenesis by upregulating the signaling of bone morphogenetic proteins

Platelet-derived growth factor receptor (PDGFR) signaling has been shown to inhibit osteogenesis. However, therapeutic efficacy of inhibiting PDGF signaling to bone regeneration in vivo and the specific mechanisms by which PDGFR signaling inhibits osteogenic differentiation remain unclear. In the present study, we examined the osteogenic effect of inhibiting PDGFR using trapidil, a PDGFR antagonist, in vivo and in vitro, and evaluated its mechanisms. A rat calvarial defect model was analyzed by micro-computed tomography and histology to determine the pro-osteogenic effect of trapidil in vivo. In addition, primary mouse calvarial osteoblast precursors were cultured in osteogenic differentiation medium with trapidil to study the mechanisms. Trapidil greatly promoted bone regeneration in a rat calvarial defect model and osteogenic differentiation of calvarial osteoblast precursors. For the mechanisms, trapidil induced phosphorylation of Smad1/5/9 and mitogen-activated protein kinase (MAPK) leading to enhance expression of Runx2, crucial transcription factor for osteogenesis. The pro-osteogenic effects of trapidil were inhibited by LDN193189, specific inhibitor of bone morphogenetic protein (BMP) receptor, ALK2 and ALK3, and by depletion of ALK3, and treatment with noggin, an antagonist of BMPs. Moreover, trapidil showed a synergistic effect with BMP2 on osteogenic differentiation. In conclusion, trapidil induced BMPR activity through upregulation of BMP signaling, leading to promoted osteogenesis in vitro and in vivo. Attenuated BMPR activity may be involved in the inhibition of osteogenesis by PDGFR signaling.