Identification of candidate neoantigens produced by fusion transcripts in human osteosarcomas.

Osteosarcomas are characterized by highly disrupted genomes. Although osteosarcomas lack common fusions, we find evidence of many tumour specific gene-gene fusion transcripts, likely due to chromosomal rearrangements and expression of transcription-induced chimeras. Most of the fusions result in out-of-frame transcripts, potentially capable of producing long novel protein sequences and a plethora of neoantigens. To identify fusions, we explored RNA-sequencing data to obtain detailed knowledge of transcribed fusions, by creating a novel program to compare fusions identified by deFuse to de novo transcripts generated by Trinity. This allowed us to confirm the deFuse results and identify unusual splicing patterns associated with fusion events. Using various existing tools combined with this custom program, we developed a pipeline for the identification of fusion transcripts applicable as targets for immunotherapy. In addition to identifying candidate neoantigens associated with fusions, we were able to use the pipeline to establish a method for measuring the frequency of fusion events, which correlated to patient outcome, as well as highlight some similarities between canine and human osteosarcomas. The results of this study of osteosarcomas underscores the numerous benefits associated with conducting a thorough analysis of fusion events within cancer samples.

Biphasic Dependence of Glioma Survival and Cell Migration on CD44 Expression Level.

While several studies link the cell-surface marker CD44 to cancer progression, conflicting results show both positive and negative correlations with increased CD44 levels. Here, we demonstrate that the survival outcomes of genetically induced glioma-bearing mice and of high-grade human glioma patients are biphasically correlated with CD44 level, with the poorest outcomes occurring at intermediate levels. Furthermore, the high-CD44-expressing mesenchymal subtype exhibited a positive trend of survival with increased CD44 level. Mouse cell migration rates in ex vivo brain slice cultures were also biphasically associated with CD44 level, with maximal migration corresponding to minimal survival. Cell simulations suggest that cell-substrate adhesiveness is sufficient to explain this biphasic migration. More generally, these results highlight the potential importance of non-monotonic relationships between survival and biomarkers associated with cancer progression.

Monomeric annexin A2 is an oxygen-regulated toll-like receptor 2 ligand and adjuvant.

BACKGROUND: Annexin A2 (ANXA2) is a pleiotropic, calcium-dependent, phospholipid-binding protein with a broad tissue distribution. It can be intracellular, membrane-bound, or secreted, and it exists as a monomer or heterotetramer. The secreted ANXA2 heterotetramer signals human and murine macrophages to produce IL-1, IL-6, and TNF-α through TLR4/MyD88- and TRIF-dependent pathways. METHODS: GL261 glioma cells were cultured in 5 % or 20 % O2. Monomeric ANXA2 (ANXA2m) was identified as a TLR2-binding protein enriched in 5 % O2 by mass spectrometry. Purified ANXA2m and ANXA2-derived peptides were added to TLR2-expressing reporter cells and immature dendritic cells (DCs) cells in vitro. ANXA2m was then mixed with chicken ovalbumin (OVA) for vaccination of TLR2 (+/+) and TLR2 (-/-) mice for subsequent quantification of antigen-specific CD8(+) T cell responses. The TLR2-binding region of ANXA2m was determined using various peptides derived from the ANXA2 amino terminus on TLR2 reporter cells and in vaccinated mice. RESULTS: ANXA2m is overexpressed by murine glioblastoma GL261 cells grown under 5 % O2, but not atmospheric 20 % O2, and acts as an adjuvant by inducing murine and human DC maturation through TLR2. ANXA2m upregulates CD80 and CD86 expression, enhances antigen cross-presentation, and induces the secretion of IL-12p70, TNF-α, and IFN-γ. The amino-terminal 15 amino acids of ANXA2m are necessary and sufficient for TLR2 binding and DC activation. CONCLUSION: This novel finding adds to the known functions of ANXA2 and suggests ways to exploit it as a vaccine adjuvant. ANXA2-antigen fusion peptides could be developed for patients as "off-the-shelf" agents containing common tumor antigens. Alternatively, they could be "personalized" and synthesized after tumor sequencing to identify immunogenic tumor-specific neo-antigens. As the amino terminal 15 amino acids of ANXA2 are required to stimulate TLR2 activity, a fusion peptide could be as short as 30 amino acids if one or two CD8 T cell epitopes are fused to the ANXA2 amino terminal portion. Future work will address the efficacy of ANXA2 peptide fusions alone and in combination with established TLR agonists to induce synergy in preclinical models of glioma as observed in other vaccines.

Effective Treatment of Established GL261 Murine Gliomas through Picornavirus Vaccination-Enhanced Tumor Antigen-Specific CD8+ T Cell Responses.

Glioblastoma (GBM) is among the most invasive and lethal of cancers, frequently infiltrating surrounding healthy tissue and giving rise to rapid recurrence. It is therefore critical to establish experimental model systems and develop therapeutic approaches that enhance anti-tumor immunity. In the current study, we have employed a newly developed murine glioma model to assess the efficacy of a novel picornavirus vaccination approach for the treatment of established tumors. The GL261-Quad system is a variation of the GL261 syngeneic glioma that has been engineered to expresses model T cell epitopes including OVA257-264. MRI revealed that both GL261 and GL261-Quad tumors display characteristic features of human gliomas such as heterogeneous gadolinium leakage and larger T2 weighted volumes. Analysis of brain-infiltrating immune cells demonstrated that GL261-Quad gliomas generate detectable CD8+ T cell responses toward the tumor-specific Kb:OVA257-264 antigen. Enhancing this response via a single intracranial or peripheral vaccination with picornavirus expressing the OVA257-264 antigen increased anti-tumor CD8+ T cells infiltrating the brain, attenuated progression of established tumors, and extended survival of treated mice. Importantly, the efficacy of the picornavirus vaccination is dependent on functional cytotoxic activity of CD8+ T cells, as the beneficial response was completely abrogated in mice lacking perforin expression. Therefore, we have developed a novel system for evaluating mechanisms of anti-tumor immunity in vivo, incorporating the GL261-Quad model, 3D volumetric MRI, and picornavirus vaccination to enhance tumor-specific cytotoxic CD8+ T cell responses and track their effectiveness at eradicating established gliomas in vivo.

Decreased affinity for efflux transporters increases brain penetrance and molecular targeting of a PI3K/mTOR inhibitor in a mouse model of glioblastoma.

BACKGROUND: Targeting drug delivery to invasive glioma cells is a particularly difficult challenge because these cells lie behind an intact blood-brain barrier (BBB) that can be observed using multimodality imaging. BBB-associated efflux transporters such as P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) influence drug distribution to these cells and may negatively impact efficacy. To test the hypothesis that efflux transporters influence brain pharmacokinetics/pharmacodynamics of molecularly targeted agents in glioma treatment, we assessed region-specific penetrance and molecular-targeting capacity for a PI3K/mTOR kinase inhibitor that has high substrate affinity for efflux transporters (GDC-0980) and an analog (GNE-317) that was purposely designed to have reduced efflux. METHODS: Brain tumor penetrance of GDC-0980 and GNE-317 was compared between FVB/n wild-type mice and Mdr1a/b(-/-)Bcrp(-/-) triple-knockout mice lacking P-gp and BCRP. C57B6/J mice bearing intracranial GL261 tumors were treated with GDC-0980, GNE-317, or vehicle to assess the targeted pharmacokinetic/pharmacodynamic effects in a glioblastoma model. RESULTS: Animals treated with GNE-317 demonstrated 3-fold greater penetrance in tumor core, rim, and normal brain compared with animals dosed with GDC-0980. Increased brain penetrance correlated with decreased staining of activated p-Akt, p-S6, and p-4EBP1 effector proteins downstream of PI3K and mTOR. CONCLUSIONS: GDC-0980 is subject to active efflux by P-gp and BCRP at the BBB, while brain penetrance of GNE-317 is independent of efflux, which translates into enhanced inhibition of PI3K/mTOR signaling. These data show that BBB efflux by P-gp and BCRP is therefore an important determinant in both brain penetrance and molecular targeting efficacy in the treatment of invasive glioma cells.

Parthenolide prodrug LC-1 slows growth of intracranial glioma.

LC-1 (also known as DMAPT or dimethylamino-parthenolide), a prodrug of parthenolide, was tested for anti-proliferative activity against glioma. LC-1 was found to have low micromolar cytotoxic activity against three glioma cell lines and was also found to be brain penetrant in healthy mice (2.1-3.0 brain-to-plasma ratio). In a syngeneic GL261 murine glioma model, LC-1 slowed tumor growth kinetics and extended the survival time of tumor-bearing mice in comparison to the vehicle control. Consequently, LC-1 represents a promising lead compound for further development as a glioma therapy.

Vaccination with dendritic cells loaded with allogeneic brain tumor cells for recurrent malignant brain tumors induces a CD4(+)IL17(+) response.

BACKGROUND: We tested the hypothesis that a novel vaccine developed from autologous dendritic cells (DC) loaded with cells from a unique allogeneic brain tumor cell line (GBM6-AD) would be well-tolerated and would generate an immune response. METHOD: Patients with recurrent primary brain tumors underwent vaccination with GBM6-AD/DC vaccine. Subjects were treated at escalating DC cell doses: 5 × 10(6) (one patient), 10 × 10(6) (one patient) and 15 × 10(6) (6 patients). Subcutaneous injections were planned for days 0, 14, 28, 42, 56, and monthly thereafter. The primary endpoint was the safety of the GBM6-AD/DC vaccination. The secondary endpoints were immune response, measured by flow cytometry, and the clinical outcome of tumor response defined by time to progression and overall survival. RESULTS: Eight patients were treated. The first three patients were treated in the dose escalation phase of the trial; the remaining five patients received the maximum dose of 15 × 10(6) DC. No dose limiting toxicity was observed. The best response per modified McDonald criteria was partial response in one patient. Flow cytometric immune profiling revealed significant differences in CD4(+)IL17(+) lymphocytes and myeloid derived suppressor cell populations between patients characterized as having stable vs. non-stable disease. CONCLUSION: This first-in-human study shows that the GBM6-AD/DC vaccine was well tolerated and was associated with an immune response in a subset of patients. No MTD was achieved in this trial. This small-scale pilot provides information for larger scale investigations into the use of this allogeneic vaccine source.

Structure-activity relationship analysis of imidazoquinolines with Toll-like receptors 7 and 8 selectivity and enhanced cytokine induction.

Toll-like receptors 7 and 8 (TLRs) have emerged as key targets in the design of small molecule adjuvants and stimulants for use in immunotherapies. This study examines the structure-activity relationship of a series of C2- and N1-substituted C7-methoxycarbonylimidazoquinolines to gain insight to the structural basis to TLR-7 and -8 selective activity. The analysis is further applied to evaluate the induction of multiple cytokines, including IL-10, IL-12, IL-1ß, TNF-α, IFN-α, and IFN-γ, using murine BMDCs and human PBMCs. The results show TLR-7/8 activity is correlated to the C2-alkyl chain length, with peak activity occurring for the butyl (TLR-7) and pentyl (TLR-8) derivatives. A similar SAR is identified in the production of IL-1ß, IL-12, and IFN-γ, which are shown to depend on both the C2-alkyl chain length and substitution to the N1-position. The compounds were also potent stimulators of IFN-α and IL-10 production but with less pronounced structure-based correlations.

CD8+ T cell-independent tumor regression induced by Fc-OX40L and therapeutic vaccination in a mouse model of glioma.

Despite the growing number of preclinical and clinical trials focused on immunotherapy for the treatment of malignant gliomas, the prognosis for this disease remains grim. Although some promising advances have been made, the immune response stimulated as a result of immunotherapeutic protocols has been inefficient at complete tumor elimination, primarily due to our lack of understanding of the necessary effector functions of the immune system. We previously demonstrated that a tumor lysate vaccine/Fc-OX40L therapy is capable of inducing enhanced survival and tumor elimination in the GL261 mouse glioma model. The following experiments were performed to determine the mechanism(s) of action of this therapy that elicits a potent antitumor immune response. The evidence subsequently outlined indicates a CD8(+) T cell-independent and CD4(+) T cell-, NK cell-, and B cell-dependent means of prolonged survival. CD8(+) T cell-independent tumor clearance is surprising considering the current focus of many cancer immunotherapy protocols. These results provide evidence for CD8(+) T cell-independent means of antitumor response and should lead to additional examination of the potential manipulation of this mechanism for future treatment strategies.

Immunogenicity of murine solid tumor models as a defining feature of in vivo behavior and response to immunotherapy.

Immune profiling has been widely used to probe mechanisms of immune escape in cancer and identify novel targets for therapy. Two emerging uses of immune signatures are to identify likely responders to immunotherapy regimens among individuals with cancer and to understand the variable responses seen among subjects with cancer in immunotherapy trials. Here, the immune profiles of 6 murine solid tumor models (CT26, 4T1, MAD109, RENCA, LLC, and B16) were correlated to tumor regression and survival in response to 2 immunotherapy regimens. Comprehensive profiles for each model were generated using quantitative reverse transcriptase polymerase chain reaction, immunohistochemistry, and flow cytometry techniques, as well as functional studies of suppressor cell populations (regulatory T cells and myeloid-derived suppressor cells), to analyze intratumoral and draining lymphoid tissues. Tumors were stratified as highly or poorly immunogenic, with highly immunogenic tumors showing a significantly greater presence of T-cell costimulatory molecules and immune suppression in the tumor microenvironment. An absence of tumor-infiltrating cytotoxic T lymphocytes and mature dendritic cells was seen across all models. Delayed tumor growth and increased survival with suppressor cell inhibition and tumor-targeted chemokine+/-dendritic cells vaccine immunotherapy were associated with high tumor immunogenicity in these models. Tumor MHC class I expression correlated with the overall tumor immunogenicity level and was a singular marker to predict immunotherapy response with these regimens. By using experimental tumor models as surrogates for human cancers, these studies demonstrate how select features of an immune profile may be utilized to identify patients most likely to respond to immunotherapy regimens.

GM-CSF promotes the immunosuppressive activity of glioma-infiltrating myeloid cells through interleukin-4 receptor-α.

Malignant gliomas are lethal cancers in the brain and heavily infiltrated by myeloid cells. Interleukin-4 receptor-α (IL-4Rα) mediates the immunosuppressive functions of myeloid cells, and polymorphisms in the IL-4Rα gene are associated with altered glioma risk and prognosis. In this study, we sought to evaluate a hypothesized causal role for IL-4Rα and myeloid suppressor cells in glioma development. In both mouse de novo gliomas and human glioblastoma cases, IL-4Rα was upregulated on glioma-infiltrating myeloid cells but not in the periphery or in normal brain. Mice genetically deficient for IL-4Rα exhibited a slower growth of glioma associated with reduced production in the glioma microenvironment of arginase, a marker of myeloid suppressor cells, which is critical for their T-cell inhibitory function. Supporting this result, investigations using bone marrow-derived myeloid cells showed that IL-4Rα mediates IL-13-induced production of arginase. Furthermore, glioma-derived myeloid cells suppressed T-cell proliferation in an IL-4Rα-dependent manner, consistent with their identification as myeloid-derived suppressor cells (MDSC). Granulocyte macrophage colony-stimulating factor (GM-CSF) plays a central role for the induction of IL-4Rα expression on myeloid cells, and we found that GM-CSF is upregulated in both human and mouse glioma microenvironments compared with normal brain or peripheral blood samples. Together, our findings establish a GM-CSF-induced mechanism of immunosuppression in the glioma microenvironment via upregulation of IL-4Rα on MDSCs.

MMuFLR: missense mutation and frameshift location reporter.

MOTIVATION: Cancer researchers seeking immunotherapy targets in cancer cells need tools to locate highly expressed proteins unique to cancer cells. Missense mutation and frameshift location reporter (MMuFLR), a Galaxy-based workflow, analyzes next-generation sequencing paired read RNA-seq output to reliably identify small frameshift mutations and missense mutations in highly expressed protein-coding genes. MMuFLR ignores known SNPs, low quality reads and poly-A/T sequences. For each frameshift and missense mutation identified, MMuFLR provides the location and sequence of the amino acid substitutions in the novel protein candidates for direct input into epitope evaluation tools. AVAILABILITY: http://toolshed.g2.bx.psu.edu/ CONTACT: rath0096@umn.edu or johns198@umn.edu SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

CD133-targeted paclitaxel delivery inhibits local tumor recurrence in a mouse model of breast cancer.

Expression of the membrane protein CD133 marks a subset of cancer cells with drug resistant phenotype and enhanced tumor initiating ability in xenotransplantation assays. Because drug resistance and tumor relapse are significant problems, approaches to eliminate these cells are urgently needed. As a step towards achieving this goal, we developed polymeric nanoparticles targeting CD133 by conjugating an anti-CD133 monoclonal antibody to nanoparticles formulated using poly(D,L lactide-co-glycolide) polymer. Nanoparticles were loaded with paclitaxel, a microtubule-stabilizing anticancer agent, as well as with 6-coumarin, a fluorescent probe. CD133-targeted nanoparticles (CD133NPs) were efficiently internalized by Caco-2 cells, which abundantly express CD133 (>9-fold higher uptake than non-targeted control nanoparticles). The effectiveness of CD133NPs in reducing tumor initiating cell (TIC) fraction was investigated using mammosphere formation and soft-agar colony formation assays. Free paclitaxel treatment was not effective in decreasing the TIC population relative to untreated control, whereas CD133NPs effectively decreased the number of mammospheres and colonies formed. In vivo studies in the MDA-MB-231 xenograft model showed that free paclitaxel was initially effective in inhibiting tumor growth but the tumors rebounded rapidly once the treatment was stopped. Tumor regrowth was significantly lower when paclitaxel was delivered through CD133NPs (tumor volume was 518.6±228 vs. 1370.9±295mm(3) for free paclitaxel at 63days; P<0.05). Our studies thus show that encapsulation of paclitaxel in CD133NPs results in a significant decrease in the TIC population and improved therapeutic efficacy compared to that with free paclitaxel treatment. These results indicate the potential of targeting anticancer therapeutics to CD133+ cells for reducing tumor recurrence.

Minicircle DNA is superior to plasmid DNA in eliciting antigen-specific CD8+ T-cell responses.

Clinical trials reveal that plasmid DNA (pDNA)-based gene delivery must be improved to realize its potential to treat human disease. Current pDNA platforms suffer from brief transgene expression, primarily due to the spread of transcriptionally repressive chromatin initially deposited on plasmid bacterial backbone sequences. Minicircle (MC) DNA lacks plasmid backbone sequences and correspondingly confers higher levels of sustained transgene expression upon delivery, accounting for its success in preclinical gene therapy models. In this study, we show for the first time that MC DNA also functions as a vaccine platform. We used a luciferase reporter transgene to demonstrate that intradermal delivery of MC DNA, relative to pDNA, resulted in significantly higher and persistent levels of luciferase expression in mouse skin. Next, we immunized mice intradermally with DNA encoding a peptide that, when presented by the appropriate major histocompatibility complex class I molecule, was recognized by endogenous CD8(+) T cells. Finally, immunization with peptide-encoding MC DNA, but not the corresponding full-length (FL) pDNA, conferred significant protection in mice challenged with Listeria monocytogenes expressing the model peptide. Together, our results suggest intradermal delivery of MC DNA may prove more efficacious for prophylaxis than traditional pDNA vaccines.

Profound impairment of adaptive immune responses by alkylating chemotherapy.

Overall, cancer vaccines have had a record of failure as an adjuvant therapy for malignancies that are treated with alkylating chemotherapy, and the contribution of standard treatment to that failure remains unclear. Vaccines aim to harness the proliferative potential of the immune system by expanding a small number of tumor-specific lymphocytes into a large number of antitumor effectors. Clinical trials are often conducted after treatment with alkylating chemotherapy, given either as standard therapy or for immunomodulatory effect. There is mounting evidence for synergy between chemotherapy and adoptive immunotherapy or vaccination against self-Ags; however, the impact of chemotherapy on lymphocytes primed against tumor neoantigens remains poorly defined. We report that clinically relevant dosages of standard alkylating chemotherapies, such as temozolomide and cyclophosphamide, significantly inhibit the proliferative abilities of lymphocytes in mice. This proliferative impairment was long-lasting and led to quantitative and qualitative defects in B and T cell responses to neoantigen vaccines. High-affinity responder lymphocytes receiving the strongest proliferative signals from vaccines experienced the greatest DNA damage responses, skewing the response toward lower-affinity responders with inferior functional characteristics. Together, these defects lead to inferior efficacy and overall survival in murine tumor models treated by neoantigen vaccines. These results suggest that clinical protocols for cancer vaccines should be designed to avoid exposing responder lymphocytes to alkylating chemotherapy.

Vaccination for invasive canine meningioma induces in situ production of antibodies capable of antibody-dependent cell-mediated cytotoxicity.

Malignant and atypical meningiomas are resistant to standard therapies and associated with poor prognosis. Despite progress in the treatment of other tumors with therapeutic vaccines, this approach has not been tested preclinically or clinically in these tumors. Spontaneous canine meningioma is a clinically meaningful but underutilized model for preclinical testing of novel strategies for aggressive human meningioma. We treated 11 meningioma-bearing dogs with surgery and vaccine immunotherapy consisting of autologous tumor cell lysate combined with toll-like receptor ligands. Therapy was well tolerated, and only one dog had tumor growth that required intervention, with a mean follow up of 585 days. IFN-γ-elaborating T cells were detected in the peripheral blood of 2 cases, but vaccine-induced tumor-reactive antibody responses developed in all dogs. Antibody responses were polyclonal, recognizing both intracellular and cell surface antigens, and HSP60 was identified as one common antigen. Tumor-reactive antibodies bound allogeneic canine and human meningiomas, showing common antigens across breed and species. Histologic analysis revealed robust infiltration of antibody-secreting plasma cells into the brain around the tumor in posttreatment compared with pretreatment samples. Tumor-reactive antibodies were capable of inducing antibody-dependent cell-mediated cytotoxicity to autologous and allogeneic tumor cells. These data show the feasibility and immunologic efficacy of vaccine immunotherapy for a large animal model of human meningioma and warrant further development toward human trials.

Vaccine injection site matters: qualitative and quantitative defects in CD8 T cells primed as a function of proximity to the tumor in a murine glioma model.

Malignant gliomas are lethal brain tumors for which novel therapies are urgently needed. In animal models, vaccination with tumor-associated Ags efficiently primes T cells to clear gliomas. In clinical trials, cancer vaccines have been less effective at priming T cells and extending survival. Generalized immune suppression in the tumor draining lymph nodes has been documented in multiple cancers. However, a systematic analysis of how vaccination at various distances from the tumor (closest to farthest) has not been reported. We investigated how the injection site chosen for vaccination dictates CD8 T cell priming and survival in an OVA-transfected murine glioma model. Glioma-bearing mice were vaccinated with Poly:ICLC plus OVA protein in the neck, hind leg, or foreleg for drainage into the cervical, inguinal, or axillary lymph nodes, respectively. OVA-specific CD8 T cell number, TCR affinity, effector function, and infiltration into the brain decreased as the vaccination site approached the tumor. These effects were dependent on the presence of the tumor, because injection site did not appreciably affect CD8 T cell priming in tumor-free mice. Our data suggest the site of vaccination can greatly impact the effectiveness of cancer vaccines. Considering that previous and ongoing clinical trials have used a variety of injection sites, vaccination site is potentially a critical aspect of study design that is being overlooked.

Function of the blood-brain barrier and restriction of drug delivery to invasive glioma cells: findings in an orthotopic rat xenograft model of glioma.

Despite aggressive treatment with radiation and chemotherapy, recurrence of glioblastoma multiforme (GBM) is inevitable. The objective of this study was to show that the blood-brain barrier (BBB), through a combination of tight junctions and active efflux transporters in the brain microvasculature, can significantly restrict delivery of molecularly targeted agents to invasive glioma cells. Transgenic mice lacking P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp) were used to study efflux of erlotinib at the BBB. A U87 rat xenograft model of GBM was used to investigate the regional distribution of erlotinib to the tumor, and brain regions surrounding the tumor. The effect of concurrent administration of elacridar on regional tumor distribution of erlotinib was evaluated. We show that erlotinib transport across an intact BBB is significantly restricted due to P-gp- and Bcrp-mediated efflux transport. We then show that the BBB is sufficiently intact in areas of brain adjacent to the tumor core to significantly restrict erlotinib delivery. Inhibition of P-gp and Bcrp by the dual inhibitor elacridar dramatically increased erlotinib delivery to the tumor core, rim, and normal brain. These results provide conclusive evidence of the impact that active efflux at the BBB has on the delivery of molecularly targeted therapy to different tumor regions in glioma. These data also support the possibility that the repeated failure of clinical trials of new drugs for gliomas may be in part due to a failure to achieve effective concentrations in invasive tumor cells that reside behind an intact BBB.

Immunologic privilege in the central nervous system and the blood-brain barrier.

The brain is in many ways an immunologically and pharmacologically privileged site. The blood-brain barrier (BBB) of the cerebrovascular endothelium and its participation in the complex structure of the neurovascular unit (NVU) restrict access of immune cells and immune mediators to the central nervous system (CNS). In pathologic conditions, very well-organized immunologic responses can develop within the CNS, raising important questions about the real nature and the intrinsic and extrinsic regulation of this immune privilege. We assess the interactions of immune cells and immune mediators with the BBB and NVU in neurologic disease, cerebrovascular disease, and intracerebral tumors. The goals of this review are to outline key scientific advances and the status of the science central to both the neuroinflammation and CNS barriers fields, and highlight the opportunities and priorities in advancing brain barriers research in the context of the larger immunology and neuroscience disciplines. This review article was developed from reports presented at the 2011 Annual Blood-Brain Barrier Consortium Meeting.