Immunosuppression is frequently associated with malignancy and is particularly severe in patients with malignant glioma. Anergy and counterproductive shifts toward T H 2 cytokine production are long-recognized T-cell defects in these patients whose etiology has remained elusive for >30 years. We show here that absolute counts of both CD4 + T cells and CD4 + CD25 + FOXP3 + CD45RO + T cells (T regs ) are greatly diminished in patients with malignant glioma, but T regs frequently represent an increased fraction of the remaining CD4 compartment. This increased T reg fraction, despite reduced counts, correlates with and is sufficient to elicit the characteristic manifestations of impaired patient T-cell responsiveness in vitro. Furthermore, T reg removal eradicates T-cell proliferative defects and reverses T H 2 cytokine shifts, allowing T cells from patients with malignant glioma to function in vitro at levels equivalent to those of normal, healthy controls. Such restored immune function may give license to physiologic antiglioma activity, as in vivo, T reg depletion proves permissive for spontaneous tumor rejection in a murine model of established intracranial glioma. These findings dramatically alter our understanding of depressed cellular immune function in patients with malignant glioma and advance a role for T regs in facilitating tumor immune evasion in the central nervous system. (Cancer Res 2006; 66(6): 3294-302)

Human cytomegalovirus (HCMV) has been described to be associated with several human malignancies, though the frequency of detection remains controversial. It is unclear whether HCMV plays an active role in malignant tumor progression or becomes reactivated under pathologic conditions that result in chronic inflammation or immunosuppression. In this study, we report on the investigation of detecting HCMV in the tumors and peripheral blood of patients with newly diagnosed glioblastoma multiforme (GBM). Using immunohistochemistry, in situ hybridization, and polymerase chain reaction amplification of viral DNA, the detection of HCMV was investigated in tumor and blood specimens from patients with GBM as well as in the peripheral blood of normal volunteers and patients undergoing craniotomy for diagnoses other than GBM. We found that a high percentage (>90%) of GBM tumors, not surrounding normal brain, are associated with HCMV nucleic acids and proteins. Furthermore, a significant proportion of patients (80%) with newly diagnosed GBM have detectable HCMV DNA in their peripheral blood, while sero-positive normal donors and other surgical patients did not exhibit detectable virus, suggesting either a systemic reactivation of HCMV within patients with GBM or shedding of viral DNA from infected tumor cells into the periphery. These results confirm the association of HCMV with malignant gliomas and demonstrate that subclinical HCMV viremia (presence of viral DNA in blood without clinical symptoms of infection) is a previously unrecognized disease spectrum in patients with GBM.

Administration of exosomes derived from mesenchymal stromal cells (MSCs) could improve some neurologic conditions by transferring functional biomolecules to recipient cells. Furthermore, exosomes from hypoxic progenitor cells exerted better therapeutic effects in organ injury through specific cargoes. However, there are no related reports about whether exosomes derived from MSCs or hypoxia-preconditioned MSCs (PC-MSCs) could prevent memory deficits in Alzheimer disease (AD). In this study, the exosomes derived from MSCs or PC-MSCs were systemically administered to transgenic APP/PS1 mice. The expression of miR-21 in MSCs was significantly increased after hypoxic treatment. Injection of exosomes from normoxic MSCs could rescue cognition and memory impairment according to results of the Morris water maze test, reduced plaque deposition, and Aβ levels in the brain; could decrease the activation of astrocytes and microglia; could down-regulate proinflammatory cytokines (TNF-α and IL-1β); and could up-regulate anti-inflammatory cytokines (IL-4 and -10) in AD mice, as well as reduce the activation of signal transducer and activator of transcription 3 (STAT3) and NF-κB. Compared to the group administered exosomes from normoxic MSCs, in the group administered exosomes from PC-MSCs, learning and memory capabilities were significantly improved; the plaque deposition and Aβ levels were lower, and expression of growth-associated protein 43, synapsin 1, and IL-10 was increased; and the levels of glial fibrillary acidic protein, ionized calcium-binding adaptor molecule 1, TNF-α, IL-1β, and activation of STAT3 and NF-κB were sharply decreased. More importantly, exosomes from PC-MSCs effectively increased the level of miR-21 in the brain of AD mice. Additionally, replenishment of miR-21 restored the cognitive deficits in APP/PS1 mice and prevented pathologic features. Taken together, these findings suggest that exosomes from PC-MSCs could improve the learning and memory capabilities of APP/PS1 mice, and that the underlying mechanism may lie in the restoration of synaptic dysfunction and regulation of inflammatory responses through regulation of miR-21.-Cui, G.-H., Wu, J., Mou, F.-F., Xie, W.-H., Wang, F.-B., Wang, Q.-L., Fang, J., Xu, Y.-W., Dong, Y.-R., Liu, J.-R., Guo, H.-D. Exosomes derived from hypoxia-preconditioned mesenchymal stromal cells ameliorate cognitive decline by rescuing synaptic dysfunction and regulating inflammatory responses in APP/PS1 mice.

Abstract:We have described recently an acetylcholinesterase (AChE) knockout mouse. While comparing the tis-

Purpose Patients with glioblastoma (GBM) have a <15 month median survival despite surgical resection, high-dose radiation and chemotherapy with temozolomide (TMZ). We previously demonstrated that targeting Cytomegalovirus (CMV) pp65 using dendritic cells (DCs) can extend survival and, in a separate study, that dose-intensified (DI) TMZ and adjuvant GM-CSF potentiates tumor-specific immune responses in patients with GBM. Here, we evaluated pp65-specific cellular responses following DI-TMZ with pp65-DCs and determined the effects on long-term progression-free survival (PFS) and overall survival (OS). Experimental Design Following standard of care, 11 patients with newly diagnosed GBM received DI-TMZ (100 mg/m2/day × 21 days per cycle) with at least three vaccines of pp65-lysosome-associated membrane glycoprotein (LAMP) mRNA-pulsed DCs admixed with GM-CSF on Day 23 ± 1 of each cycle. Thereafter, monthly DI-TMZ cycles and pp65-DCs were continued if patients had not progressed. Results Following DI-TMZ cycle 1 and three doses of pp65-DCs, pp65 cellular responses significantly increased. After DI-TMZ, both the proportion and proliferation of regulatory T-cells (TRegs) increased and remained elevated with serial DI-TMZ cycles. Median PFS and OS were 25.3 months (CI95: 11.0-∞) and 41.1 months (CI95: 21.6-∞), exceeding survival using recursive partitioning analysis and matched historical controls. Four patients remained progression-free at 59 to 64 months from diagnosis. No known prognostic factors (age, KPS, IDH-1/2 mutation, and MGMT promoter methylation) predicted more favorable outcomes for the patients in this cohort. Conclusions Despite increased TReg proportions following DI-TMZ, patients receiving pp65-DCs showed long-term PFS and OS, confirming prior studies targeting CMV in GBM.