Cellular Therapy for Children with Central Nervous System Tumors: Mining and Mapping the Correlative Data.

PURPOSE OF REVIEW: Correlative studies should leverage clinical trial frameworks to conduct biospecimen analyses that provide insight into the bioactivity of the intervention and facilitate iteration toward future trials that further improve patient outcomes. In pediatric cellular immunotherapy trials, correlative studies enable deeper understanding of T cell mobilization, durability of immune activation, patterns of toxicity, and early detection of treatment response. Here, we review the correlative science in adoptive cell therapy (ACT) for childhood central nervous system (CNS) tumors, with a focus on existing chimeric antigen receptor (CAR) and T cell receptor (TCR)-expressing T cell therapies. RECENT FINDINGS: We highlight long-standing and more recently understood challenges for effective alignment of correlative data and offer practical considerations for current and future approaches to multi-omic analysis of serial tumor, serum, and cerebrospinal fluid (CSF) biospecimens. We highlight the preliminary success in collecting serial cytokine and proteomics from patients with CNS tumors on ACT clinical trials.

Correction to: Comparative multidimensional molecular analyses of pediatric diffuse intrinsic pontine glioma reveals distinct molecular subtypes.

The original version of this article unfortunately contained a mistake.

A Proteogenomic Approach to Understanding MYC Function in Metastatic Medulloblastoma Tumors.

Brain tumors are the leading cause of cancer-related deaths in children, and medulloblastoma is the most prevalent malignant childhood/pediatric brain tumor. Providing effective treatment for these cancers, with minimal damage to the still-developing brain, remains one of the greatest challenges faced by clinicians. Understanding the diverse events driving tumor formation, maintenance, progression, and recurrence is necessary for identifying novel targeted therapeutics and improving survival of patients with this disease. Genomic copy number alteration data, together with clinical studies, identifies c-MYC amplification as an important risk factor associated with the most aggressive forms of medulloblastoma with marked metastatic potential. Yet despite this, very little is known regarding the impact of such genomic abnormalities upon the functional biology of the tumor cell. We discuss here how recent advances in quantitative proteomic techniques are now providing new insights into the functional biology of these aggressive tumors, as illustrated by the use of proteomics to bridge the gap between the genotype and phenotype in the case of c-MYC-amplified/associated medulloblastoma. These integrated proteogenomic approaches now provide a new platform for understanding cancer biology by providing a functional context to frame genomic abnormalities.

Phase I and pharmacokinetic trial of PTC299 in pediatric patients with refractory or recurrent central nervous system tumors: a PBTC study.

PTC299 is a novel, orally-bioavailable small molecule that selectively inhibits vascular endothelial growth factor receptor protein synthesis at the post-transcriptional level. Based on promising preclinical results, we conducted a pediatric phase I study to estimate the maximum tolerated dose, describe dose-limiting toxicities (DLT) and characterize the pharmacokinetic profile of PTC299 in children with recurrent CNS tumors. PTC299 was administered orally twice or three times daily, depending on the regimen. Four regimens were evaluated using the rolling 6 design, starting with 1.2 mg/kg/dose twice daily and escalating to 2 mg/kg/dose three times daily. Pharmacokinetic studies were performed during the first two courses. Twenty-seven children (14 male, median age 11.2, range 5.5-21 years) with recurrent brain tumors were treated; 21 were fully evaluable for toxicity assessment. Therapy was well-tolerated, and the only DLT was grade 3 hyponatremia. Grade three and grade four toxicities were uncommon in subsequent cycles. Median AUC0-Tlast values at the 2 mg/kg were similar to those observed in adults. The study was terminated while patients were being treated at the highest planned dose, due to hepatotoxicity encountered in the ongoing adult phase I studies. No complete or partial responses were observed. Two patients with low-grade gliomas were noted to have minor responses, and at the time of the study's closure, 5 children with low-grade gliomas had been on therapy for 8 or more courses (range 8-16). PTC299 was well-tolerated at the highest dose level tested (2 mg/kg/dose TID) in children with recurrent brain tumors and prolonged disease stabilization was seen in children with low-grade gliomas.

Preclinical magnetic resonance imaging and systems biology in cancer research: current applications and challenges.

Biologically accurate mouse models of human cancer have become important tools for the study of human disease. The anatomical location of various target organs, such as brain, pancreas, and prostate, makes determination of disease status difficult. Imaging modalities, such as magnetic resonance imaging, can greatly enhance diagnosis, and longitudinal imaging of tumor progression is an important source of experimental data. Even in models where the tumors arise in areas that permit visual determination of tumorigenesis, longitudinal anatomical and functional imaging can enhance the scope of studies by facilitating the assessment of biological alterations, (such as changes in angiogenesis, metabolism, cellular invasion) as well as tissue perfusion and diffusion. One of the challenges in preclinical imaging is the development of infrastructural platforms required for integrating in vivo imaging and therapeutic response data with ex vivo pathological and molecular data using a more systems-based multiscale modeling approach. Further challenges exist in integrating these data for computational modeling to better understand the pathobiology of cancer and to better affect its cure. We review the current applications of preclinical imaging and discuss the implications of applying functional imaging to visualize cancer progression and treatment. Finally, we provide new data from an ongoing preclinical drug study demonstrating how multiscale modeling can lead to a more comprehensive understanding of cancer biology and therapy.

Comparative multidimensional molecular analyses of pediatric diffuse intrinsic pontine glioma reveals distinct molecular subtypes.

Diffuse intrinsic pontine glioma (DIPG) is a highly morbid form of pediatric brainstem glioma. Here, we present the first comprehensive protein, mRNA, and methylation profiles of fresh-frozen DIPG specimens (n = 14), normal brain tissue (n = 10), and other pediatric brain tumors (n = 17). Protein profiling identified 2,305 unique proteins indicating distinct DIPG protein expression patterns compared to other pediatric brain tumors. Western blot and immunohistochemistry validated upregulation of Clusterin (CLU), Elongation Factor 2 (EF2), and Talin-1 (TLN1) in DIPGs studied. Comparisons to mRNA expression profiles generated from tumor and adjacent normal brain tissue indicated two DIPG subgroups, characterized by upregulation of Myc (N-Myc) or Hedgehog (Hh) signaling. We validated upregulation of PTCH, a membrane receptor in the Hh signaling pathway, in a subgroup of DIPG specimens. DNA methylation analysis indicated global hypomethylation of DIPG compared to adjacent normal tissue specimens, with differential methylation of 24 genes involved in Hh and Myc pathways, correlating with protein and mRNA expression patterns. Sequencing analysis showed c.83A>T mutations in the H3F3A or HIST1H3B gene in 77 % of our DIPG cohort. Supervised analysis revealed a unique methylation pattern in mutated specimens compared to the wild-type DIPG samples. This study presents the first comprehensive multidimensional protein, mRNA, and methylation profiling of pediatric brain tumor specimens, detecting the presence of two subgroups within our DIPG cohort. This multidimensional analysis of DIPG provides increased analytical power to more fully explore molecular signatures of DIPGs, with implications for evaluating potential molecular subtypes and biomarker discovery for assessing response to therapy.

The Open Pediatric Cancer Project.

Background: In 2019, the Open Pediatric Brain Tumor Atlas (OpenPBTA) was created as a global, collaborative open-science initiative to genomically characterize 1,074 pediatric brain tumors and 22 patient-derived cell lines. Here, we extend the OpenPBTA to create the Open Pediatric Cancer (OpenPedCan) Project, a harmonized open-source multi-omic dataset from 6,112 pediatric cancer patients with 7,096 tumor events across more than 100 histologies. Combined with RNA-Seq from the Genotype-Tissue Expression (GTEx) and The Cancer Genome Atlas (TCGA), OpenPedCan contains nearly 48,000 total biospecimens (24,002 tumor and 23,893 normal specimens). Findings: We utilized Gabriella Miller Kids First (GMKF) workflows to harmonize WGS, WXS, RNA-seq, and Targeted Sequencing datasets to include somatic SNVs, InDels, CNVs, SVs, RNA expression, fusions, and splice variants. We integrated summarized CPTAC whole cell proteomics and phospho-proteomics data, miRNA-Seq data, and have developed a methylation array harmonization workflow to include m-values, beta-vales, and copy number calls. OpenPedCan contains reproducible, dockerized workflows in GitHub, CAVATICA, and Amazon Web Services (AWS) to deliver harmonized and processed data from over 60 scalable modules which can be leveraged both locally and on AWS. The processed data are released in a versioned manner and accessible through CAVATICA or AWS S3 download (from GitHub), and queryable through PedcBioPortal and the NCI's pediatric Molecular Targets Platform. Notably, we have expanded PBTA molecular subtyping to include methylation information to align with the WHO 2021 Central Nervous System Tumor classifications, allowing us to create research-grade integrated diagnoses for these tumors. Conclusions: OpenPedCan data and its reproducible analysis module framework are openly available and can be utilized and/or adapted by researchers to accelerate discovery, validation, and clinical translation.

The receptor tyrosine kinase EphA2 is a direct target gene of hypermethylated in cancer 1 (HIC1).

The tumor suppressor gene hypermethylated in cancer 1 (HIC1), which encodes a transcriptional repressor, is epigenetically silenced in many human tumors. Here, we show that ectopic expression of HIC1 in the highly malignant MDA-MB-231 breast cancer cell line severely impairs cell proliferation, migration, and invasion in vitro. In parallel, infection of breast cancer cell lines with a retrovirus expressing HIC1 also induces decreased mRNA and protein expression of the tyrosine kinase receptor EphA2. Moreover, chromatin immunoprecipitation (ChIP) and sequential ChIP experiments demonstrate that endogenous HIC1 proteins are bound, together with the MTA1 corepressor, to the EphA2 promoter in WI38 cells. Taken together, our results identify EphA2 as a new direct target gene of HIC1. Finally, we observe that inactivation of endogenous HIC1 through RNA interference in normal breast epithelial cells results in the up-regulation of EphA2 and is correlated with increased cellular migration. To conclude, our results involve the tumor suppressor HIC1 in the transcriptional regulation of the tyrosine kinase receptor EphA2, whose ligand ephrin-A1 is also a HIC1 target gene. Thus, loss of the regulation of this Eph pathway through HIC1 epigenetic silencing could be an important mechanism in the pathogenesis of epithelial cancers.

Loss of Hypermethylated in Cancer 1 (HIC1) in breast cancer cells contributes to stress-induced migration and invasion through ß-2 adrenergic receptor (ADRB2) misregulation.

The transcriptional repressor HIC1 (Hypermethylated in Cancer 1) is a tumor suppressor gene inactivated in many human cancers including breast carcinomas. In this study, we show that HIC1 is a direct transcriptional repressor of ß-2 adrenergic receptor (ADRB2). Through promoter luciferase activity, chromatin immunoprecipitation (ChIP) and sequential ChIP experiments, we demonstrate that ADRB2 is a direct target gene of HIC1, endogenously in WI-38 cells and following HIC1 re-expression in breast cancer cells. Agonist-mediated stimulation of ADRB2 increases the migration and invasion of highly malignant MDA-MB-231 breast cancer cells but these effects are abolished following HIC1 re-expression or specific down-regulation of ADRB2 by siRNA treatment. Our results suggest that early inactivation of HIC1 in breast carcinomas could predispose to stress-induced metastasis through up-regulation of the ß-2 adrenergic receptor.

The Reelin receptors ApoER2 and VLDLR are direct target genes of HIC1 (Hypermethylated In Cancer 1).

The tumor suppressor gene HIC1 (Hypermethylated In Cancer 1) is located in 17p13.3 a region frequently hypermethylated or deleted in tumors and in a contiguous-gene syndrome, the Miller-Dieker syndrome which includes classical lissencephaly (smooth brain) and severe developmental defects. HIC1 encodes a transcriptional repressor involved in the regulation of growth control, DNA damage response and cell migration properties. We previously demonstrated that the membrane-associated G-protein-coupled receptors CXCR7, ADRB2 and the tyrosine kinase receptor EphA2 are direct target genes of HIC1. Here we show that ectopic expression of HIC1 in U2OS and MDA-MB-231 cell lines decreases expression of the ApoER2 and VLDLR genes, encoding two canonical tyrosine kinase receptors for Reelin. Conversely, knock-down of endogenous HIC1 in BJ-Tert normal human fibroblasts through RNA interference results in the up-regulation of these two Reelin receptors. Finally, through chromatin immunoprecipitation (ChIP) in BJ-Tert fibroblasts, we demonstrate that HIC1 is a direct transcriptional repressor of ApoER2 and VLDLR. These data provide evidence that HIC1 is a new regulator of the Reelin pathway which is essential for the proper migration of neuronal precursors during the normal development of the cerebral cortex, of Purkinje cells in the cerebellum and of mammary epithelial cells. Deregulation of this pathway through HIC1 inactivation or deletion may contribute to its role in tumor promotion. Moreover, HIC1, through the direct transcriptional repression of ATOH1 and the Reelin receptors ApoER2 and VLDLR, could play an essential role in normal cerebellar development.

Long-term efficacy and toxicity of bevacizumab-based therapy in children with recurrent low-grade gliomas.

BACKGROUND: Because definitive resection or radiotherapy for pediatric low-grade gliomas (LGGs) may be associated with severe and permanent adverse effects, medical management has taken a significant role. Bevacizumab-based therapy has demonstrated encouraging responses; however, longer-term toxicity, response durability and alternative dosing regimens have not been evaluated. PROCEDURE: This was a retrospective review of children with multiply recurrent, progressive LGGs treated with bevacizumab-based therapy and followed for at least 12 months after treatment completion. Toxicity was uniformly graded and imaging was centrally reviewed. RESULTS: All fourteen patients had failed at least two prior treatment regimens; six had dissemination. Patients received initial bevacizumab-based therapy at a median age of 5.3 years (range, 1-12 years). Median treatment duration was 12 months (range, 1-24 months). 12 patients had an objective response; 2 had stable disease. Median time to maximum response was 9 weeks (range, 7-17 weeks). No patients progressed on therapy, although 13/14 progressed after stopping bevacizumab at a median of 5 months. Four patients were re-treated with bevacizumab and all again responded or stabilized. Alternative dosing strategies were effective, including bevacizumab monotherapy and prolonging the dosing interval to 3 weeks. High-grade bevacizumab-related toxicities consisted of grade 3 proteinuria (n = 2), primary inflammatory arthritis (n = 1), and somnolence (n = 1). Toxicities resolved within 6 months of treatment cessation except one case of hypertension. CONCLUSIONS: Bevacizumab-based therapy is successful at inducing rapid LGG response. Patients progressing off-therapy may be successfully re-treated with bevacizumab. Nearly all tumors progress once treatment is discontinued. Toxicities are not insignificant but usually reversible.

OpenPBTA: The Open Pediatric Brain Tumor Atlas.

Pediatric brain and spinal cancers are collectively the leading disease-related cause of death in children; thus, we urgently need curative therapeutic strategies for these tumors. To accelerate such discoveries, the Children's Brain Tumor Network (CBTN) and Pacific Pediatric Neuro-Oncology Consortium (PNOC) created a systematic process for tumor biobanking, model generation, and sequencing with immediate access to harmonized data. We leverage these data to establish OpenPBTA, an open collaborative project with over 40 scalable analysis modules that genomically characterize 1,074 pediatric brain tumors. Transcriptomic classification reveals universal TP53 dysregulation in mismatch repair-deficient hypermutant high-grade gliomas and TP53 loss as a significant marker for poor overall survival in ependymomas and H3 K28-mutant diffuse midline gliomas. Already being actively applied to other pediatric cancers and PNOC molecular tumor board decision-making, OpenPBTA is an invaluable resource to the pediatric oncology community.

Targeted gene expression profiling predicts meningioma outcomes and radiotherapy responses.

Surgery is the mainstay of treatment for meningioma, the most common primary intracranial tumor, but improvements in meningioma risk stratification are needed and indications for postoperative radiotherapy are controversial. Here we develop a targeted gene expression biomarker that predicts meningioma outcomes and radiotherapy responses. Using a discovery cohort of 173 meningiomas, we developed a 34-gene expression risk score and performed clinical and analytical validation of this biomarker on independent meningiomas from 12 institutions across 3 continents (N = 1,856), including 103 meningiomas from a prospective clinical trial. The gene expression biomarker improved discrimination of outcomes compared with all other systems tested (N = 9) in the clinical validation cohort for local recurrence (5-year area under the curve (AUC) 0.81) and overall survival (5-year AUC 0.80). The increase in AUC compared with the standard of care, World Health Organization 2021 grade, was 0.11 for local recurrence (95% confidence interval 0.07 to 0.17, P < 0.001). The gene expression biomarker identified meningiomas benefiting from postoperative radiotherapy (hazard ratio 0.54, 95% confidence interval 0.37 to 0.78, P = 0.0001) and suggested postoperative management could be refined for 29.8% of patients. In sum, our results identify a targeted gene expression biomarker that improves discrimination of meningioma outcomes, including prediction of postoperative radiotherapy responses.

Advances in the proteomic investigation of the cell secretome.

Studies of the cell secretome have greatly increased in recent years owing to improvements in proteomic platforms, mass spectrometry instrumentation and to the increased interaction between analytical chemists, biologists and clinicians. Several secretome studies have been implemented in different areas of research, leading to the generation of a valuable secretome catalogs. Secreted proteins continue to be an important source of biomarkers and therapeutic target discovery and are equally valuable in the field of microbiology. Several discoveries have been achieved in vitro using cell culture systems, ex vivo using human tissue specimens and in vivo using animal models. In this review, some of the most recent advances in secretome studies and the fields that have benefited the most from this evolving technology are highlighted.

OLIG2 Is a Determinant for the Relapse of MYC-Amplified Medulloblastoma.

PURPOSE: Patients with MYC-amplified medulloblastoma (MB) have poor prognosis and frequently develop recurrence, thus new therapeutic approaches to prevent recurrence are needed. EXPERIMENTAL DESIGN: We evaluated OLIG2 expression in a panel of mouse Myc-driven MB tumors, patient MB samples, and patient-derived xenograft (PDX) tumors and analyzed radiation sensitivity in OLIG2-high and OLIG2-low tumors in PDX lines. We assessed the effect of inhibition of OLIG2 by OLIG2-CRISPR or the small molecule inhibitor CT-179 combined with radiotherapy on tumor progression in PDX models. RESULTS: We found that MYC-associated MB can be stratified into OLIG2-high and OLIG2-low tumors based on OLIG2 protein expression. In MYC-amplified MB PDX models, OLIG2-low tumors were sensitive to radiation and rarely relapsed, whereas OLIG2-high tumors were resistant to radiation and consistently developed recurrence. In OLIG2-high tumors, irradiation eliminated the bulk of tumor cells; however, a small number of tumor cells comprising OLIG2- tumor cells and rare OLIG2+ tumor cells remained in the cerebellar tumor bed when examined immediately post-irradiation. All animals harboring residual-resistant tumor cells developed relapse. The relapsed tumors mirrored the cellular composition of the primary tumors with enriched OLIG2 expression. Further studies demonstrated that OLIG2 was essential for recurrence, as OLIG2 disruption with CRISPR-mediated deletion or with the small molecule inhibitor CT-179 prevented recurrence from the residual radioresistant tumor cells. CONCLUSIONS: Our studies reveal that OLIG2 is a biomarker and an effective therapeutic target in a high-risk subset of MYC-amplified MB, and OLIG2 inhibitor combined with radiotherapy represents a novel effective approach for treating this devastating disease.

Proteomic profiling of cerebrospinal fluid identifies prostaglandin D2 synthase as a putative biomarker for pediatric medulloblastoma: A pediatric brain tumor consortium study.

The aims of this study were to demonstrate the feasibility of centrally collecting and processing high-quality cerebrospinal fluid (CSF) samples for proteomic studies within a multi-center consortium and to identify putative biomarkers for medulloblastoma in CSF. We used 2-DE to investigate the CSF proteome from 33 children with medulloblastoma and compared it against the CSF proteome from 25 age-matched controls. Protein spots were subsequently identified by a combination of in-gel tryptic digestion and MALDI-TOF TOF MS analysis. On average, 160 protein spots were detected by 2-DE and 76 protein spots corresponding to 25 unique proteins were identified using MALDI-TOF. Levels of prostaglandin D2 synthase (PGD2S) were found to be six-fold decreased in the tumor samples versus control samples (p<0.00001). These data were further validated using ELISA. Close examination of PGD2S spots revealed the presence of complex sialylated carbohydrates at residues Asn(78) and Asn(87) . Total PGD2S levels are reduced six-fold in the CSF of children with medulloblastoma most likely representing a host response to the presence of the tumor. In addition, our results demonstrate the feasibility of performing proteomic studies on CSF samples collected from patients at multiple institutions within the consortium setting.

Isolated extracranial recurrence of anaplastic ependymoma.

Anaplastic ependymoma is a malignant glial tumor thought to arise from radial glial cells of the ventricular zone. Because ependymoma is frequently encountered within ventricular spaces, they are prone to leptomeningeal dissemination. Metastatic extracranial ependymoma has been reported, but in the context of progressive intracranial disease. We report on a boy who developed isolated extracranial recurrence of his anaplastic ependymoma, initially at the scalp and later metastases to cervical lymph nodes. The location of tumor recurrence proximate to the surgical site suggested surgical seeding. This case demonstrates an unusual site of recurrence of anaplastic ependymoma and highlights a previously underappreciated surgical complication.

CD34⁺ collection efficiency as a function of blood volumes processed in pediatric autologous peripheral blood stem cell collection.

PURPOSE: To characterize the relationship between CD34(+) collection efficiency and blood volumes processed in pediatric patients undergoing autologous peripheral blood stem cell (PBSC) collection. METHODS: Retrospective 8-year (2001-2009) study of pediatric patients (n = 79) with neuroblastoma and central nervous system (CNS) tumors undergoing first day of autologous PBSC harvest using MNC program on the COBE Spectra (Caridian BCT, Lakewood, CO) was performed. Patients undergoing 0 to 2.9 BV (standard volume), 3 to 6 BV (large volume), and greater than 6 BV (ultra large volume) harvests were evaluated for CD34(+) collection efficiency, diagnosis (neuroblastoma vs. nonneuroblastoma), disease type (primary vs. relapse), mobilization regimen, granulocyte colony stimulating factor (GCSF) dose, and apheresis complications. RESULTS: CD34(+) collection efficiencies (CE) for neuroblastoma patients were 67%, 50%, and 53% for standard (n = 14), large (n = 9), and ultra large (n = 5) volume harvests, respectively. Similarly, patients with nonneuroblastoma diagnoses had CD34(+) CE of 63%, 55%, and 65% for low (n = 19), large (n = 27), and ultra large (n = 5) volume harvests, respectively. Weight, granulocyte colony stimulating factor (G-CSF) stimulation, type of mobilization, and apheresis complications (normalized by run time) were similar between the standard, large, and ultra large volume groups in patients with either neuroblastoma or nonneuroblastoma diagnoses. CONCLUSIONS: CD34(+) collection efficiency in pediatric autologous PBSC collection on the first day of harvest does not decrease with higher numbers of blood volumes processed in patients with either neuroblastoma or nonneuroblastoma primary disease. These results indirectly indicate bone marrow CD34(+) cell mobilization occurs with longer apheresis procedures in pediatric patients.

Proteogenomic discovery of neoantigens facilitates personalized multi-antigen targeted T cell immunotherapy for brain tumors.

Neoantigen discovery in pediatric brain tumors is hampered by their low mutational burden and scant tissue availability. Here we develop a proteogenomic approach combining tumor DNA/RNA sequencing and mass spectrometry proteomics to identify tumor-restricted (neoantigen) peptides arising from multiple genomic aberrations to generate a highly target-specific, autologous, personalized T cell immunotherapy. Our data indicate that aberrant splice junctions are the primary source of neoantigens in medulloblastoma, a common pediatric brain tumor. Proteogenomically identified tumor-specific peptides are immunogenic and generate MHC II-based T cell responses. Moreover, polyclonal and polyfunctional T cells specific for tumor-specific peptides effectively eliminate tumor cells in vitro. Targeting tumor-specific antigens obviates the issue of central immune tolerance while potentially providing a safety margin favoring combination with other immune-activating therapies. These findings demonstrate the proteogenomic discovery of immunogenic tumor-specific peptides and lay the groundwork for personalized targeted T cell therapies for children with brain tumors.

Scavenger chemokine (CXC motif) receptor 7 (CXCR7) is a direct target gene of HIC1 (hypermethylated in cancer 1).

The tumor suppressor gene HIC1 (Hypermethylated in Cancer 1) that is epigenetically silenced in many human tumors and is essential for mammalian development encodes a sequence-specific transcriptional repressor. The few genes that have been reported to be directly regulated by HIC1 include ATOH1, FGFBP1, SIRT1, and E2F1. HIC1 is thus involved in the complex regulatory loops modulating p53-dependent and E2F1-dependent cell survival and stress responses. We performed genome-wide expression profiling analyses to identify new HIC1 target genes, using HIC1-deficient U2OS human osteosarcoma cells infected with adenoviruses expressing either HIC1 or GFP as a negative control. These studies identified several putative direct target genes, including CXCR7, a G-protein-coupled receptor recently identified as a scavenger receptor for the chemokine SDF-1/CXCL12. CXCR7 is highly expressed in human breast, lung, and prostate cancers. Using quantitative reverse transcription-PCR analyses, we demonstrated that CXCR7 was repressed in U2OS cells overexpressing HIC1. Inversely, inactivation of endogenous HIC1 by RNA interference in normal human WI38 fibroblasts results in up-regulation of CXCR7 and SIRT1. In silico analyses followed by deletion studies and luciferase reporter assays identified a functional and phylogenetically conserved HIC1-responsive element in the human CXCR7 promoter. Moreover, chromatin immunoprecipitation (ChIP) and ChIP upon ChIP experiments demonstrated that endogenous HIC1 proteins are bound together with the C-terminal binding protein corepressor to the CXCR7 and SIRT1 promoters in WI38 cells. Taken together, our results implicate the tumor suppressor HIC1 in the transcriptional regulation of the chemokine receptor CXCR7, a key player in the promotion of tumorigenesis in a wide variety of cell types.