Combination therapy with Treg and mesenchymal stromal cells enhances potency and attenuation of inflammation after traumatic brain injury compared to monotherapy.

The inflammatory response after traumatic brain injury (TBI) can lead to significant secondary brain injury and chronic inflammation within the central nervous system. Cell therapies, including mesenchymal stromal cells (MSC), have led to improvements in animal models of TBI and are under investigation in human trials. One potential mechanism for the therapeutic potential of MSC is their ability to augment the endogenous response of immune suppressive regulatory T cells (Treg). We have recently shown that infusion of human cord blood Treg decreased chronic microgliosis after TBI and altered the systemic immune response in a rodent model. These cells likely use both overlapping and distinct mechanisms to modulate the immune system; therefore, combining Treg and MSC as a combination therapy may confer therapeutic benefit over either monotherapy. However, investigation of Treg + MSC combination therapy in TBI is lacking. In this study, we compared the ability MSC + Treg combination therapy, as well as MSC and Treg monotherapies, to inhibit the neuroinflammatory response to TBI in vivo and in vitro. Treg + MSC combination therapy demonstrated increased potency to reduce the neuro- and peripheral inflammatory response compared to monotherapy; furthermore, the timing of infusion proved to be a significant variable in the efficacy of both MSC monotherapy and Treg + MSC combination therapy in vivo and in vitro.

Variable expression of CXCR4 in molecular subtypes of infiltrating gliomas.

AIM: Glioblastomas (GBMs) and diffuse intrinsic pontine gliomas (DIPGs) are infiltrating gliomas with poor prognosis. CXCR4 has been linked to glioma cell invasion, survival, proliferation, and angiogenesis. This study aimed to evaluate the expression of CXCR4 in molecular subtypes of adult and pediatric infiltrating gliomas. MATERIALS AND METHODS: We evaluated the expression of CXCR4 in 21 DIPGs and 44 adult infiltrating gliomas (25 GBM, 8 astrocytomas, and 11 oligodendrogliomas) by immunohistochemistry. Mutations in 315 cancer genes and rearrangements in 28 genes were evaluated by next-generation sequencing. RESULTS: CXCR4 was expressed in -DIPGs and adult infiltrating gliomas in tumor cells (28.6% and 5.6%, respectively) and endothelial cells (14.3% and 19.4%?, respectively). In adult gliomas, there was a correlation between CXCR4 expression and mutations in EGFR, PIK3CA, TERT promoter, and CDKN2A/B loss. In contrast, CXCR4 expression was not detected in IDH1/IDH2 mutant gliomas. These associations were confirmed using The Cancer Genome Atlas (TCGA) database. CONCLUSION: CXCR4 is expressed in a subset of DIPGs and GBMs, but it is not expressed in astrocytomas or oligodendrogliomas. CXCR4 expression is variable and it is influenced by tumor genomic alterations. It is important to consider CXCR4 expression in clinical trials that evaluate the efficacy of CXCR4 inhibitors in the treatment of gliomas.

Cerebral Radiation Necrosis: Incidence, Pathogenesis, Diagnostic Challenges, and Future Opportunities.

PURPOSE OF REVIEW: Cerebral radiation necrosis (CRN) is a major dose-limiting adverse event of radiotherapy. The incidence rate of RN varies with the radiotherapy modality, total dose, dose fractionation, and the nature of the lesion being targeted. In addition to these known and controllable features, there is a stochastic component to the occurrence of CRN-the genetic profile of the host or the lesion and their role in the development of CRN. RECENT FINDINGS: Recent studies provide some insight into the genetic mechanisms underlying radiation-induced brain injury. In addition to these incompletely understood host factors, the diagnostic criteria for CRN using structural and functional imaging are also not clear, though multiple structural and functional imaging modalities exist, a combination of which may prove to be the ideal diagnostic imaging approach. As the utilization of novel molecular therapies and immunotherapy increases, the incidence of CNR is expected to increase and its diagnosis will become more challenging. Tissue biopsies can be insensitive and suffer from sampling biases and procedural risks. Liquid biopsies represent a promising, accurate, and non-invasive diagnostic strategy, though this modality is currently in its infancy. A better understanding of the pathogenesis of CRN will expand and optimize the diagnosis and management of CRN by better utilizing existing treatment options including bevacizumab, pentoxifylline, hyperbaric oxygen therapy, and laser interstitial thermal therapy.

Characterization of genomic alterations in primary central nervous system lymphomas.

PURPOSE: Primary central nervous system lymphoma (PCNSL) is a non-Hodgkin lymphoma that affects the central nervous system (CNS). Although previous studies have reported the most common mutated genes in PCNSL, including MYD88 and CD79b, our understanding of genetic characterizations in primary CNS lymphomas is limited. The aim of this study was to perform a retrospective analysis investigating the most frequent mutation types, and their frequency, in PCNSL. METHODS: Fifteen patients with a diagnosis of PCNSL from our institution were analyzed for mutations in 406 genes and rearrangements in 31 genes by next generation sequencing (NGS). RESULTS: Missense mutations were identified as the most common mutation type (32%) followed by frame shift mutations (23%). The highest mutation rate was reported in the MYD88 (33.3%), CDKN2A/B (33.3%), and TP53 (26.7%) genes. Intermediate tumor mutation burden (TMB) and high TMB was detected in 13.3% and 26.7% of PCNSL, respectively. The most frequent gene rearrangement involved the IGH-BCL6 genes (20%). CONCLUSIONS: This study shows the most common genetic alterations in PCNSL as determined by a commercial next generation sequencing assay. MYD88 and CD79b are frequently mutated in PCNSL, IGH-BCL6 is the most frequent gene rearrangement and approximately 1/4 of cases show a high TMB. Mutations in multiple genes, in addition to high TMB and gene rearrangements, highlights the complex molecular heterogeneity of PCNSL. Knowledge about genetic alterations in PCNSL can inform the development of novel targets for diagnosis and treatment.

Human-derived Treg and MSC combination therapy may augment immunosuppressive potency in vitro, but did not improve blood brain barrier integrity in an experimental rat traumatic brain injury model.

Traumatic brain injury (TBI) causes both physical disruption of the blood brain barrier (BBB) and altered immune responses that can lead to significant secondary brain injury and chronic inflammation within the central nervous system (CNS). Cell therapies, including mesenchymal stromal cells (MSC), have been shown to restore BBB integrity and augment endogenous splenic regulatory T cells (Treg), a subset of CD4+ T cells that function to regulate immune responses and prevent autoimmunity. We have recently shown that infusion of human cord blood-derived Treg decreased neuroinflammation after TBI in vivo and in vitro. However, while both cells have demonstrated anti-inflammatory and regenerative potential, they likely utilize differing, although potentially overlapping, mechanisms. Furthermore, studies investigating these two cell types together, as a combination therapy, are lacking. In this study, we compared the ability of Treg+MSC combination therapy, as well as MSC and Treg monotherapies, to improve BBB permeability in vivo and suppress inflammation in vitro. While Treg+MSC combination did not significantly augment potency in vivo, our in vitro data demonstrates that combination therapy may augment therapeutic potency and immunosuppressive potential compared to Treg or MSC monotherapy.

Glioma and temozolomide induced alterations in gut microbiome.

The gut microbiome is fundamental in neurogenesis processes. Alterations in microbial constituents promote inflammation and immunosuppression. Recently, in immune-oncology, specific microbial taxa have been described to enhance the effects of therapeutic modalities. However, the effects of microbial dysbiosis on glioma are still unknown. The aim of this study was to explore the effects of glioma development and Temozolomide (TMZ) on fecal microbiome in mice and humans. C57BL/6 mice were implanted with GL261/Sham and given TMZ/Saline. Fecal samples were collected longitudinally and analyzed by 16S rRNA sequencing. Fecal samples were collected from healthy controls as well as glioma patients at diagnosis, before and after chemoradiation. Compared to healthy controls, mice and glioma patients demonstrated significant differences in beta diversity, Firmicutes/Bacteroides (F/B) ratio, and increase of Verrucomicrobia phylum and Akkermansia genus. These changes were not observed following TMZ in mice. TMZ treatment in the non-tumor bearing mouse-model diminished the F/B ratio, increase Muribaculaceae family and decrease Ruminococcaceae family. Nevertheless, there were no changes in Verrucomicrobia/Akkermansia. Glioma development leads to gut dysbiosis in a mouse-model, which was not observed in the setting of TMZ. These findings seem translational to humans and warrant further study.

Circulating tumour DNA, microRNA and metabolites in cerebrospinal fluid as biomarkers for central nervous system malignancies.

Central nervous system (CNS) malignancies can be difficult to diagnose and many do not respond satisfactorily to existing therapies. Monitoring patients with CNS malignancies for treatment response and tumour recurrence can be challenging because of the difficulty and risks of brain biopsies, and the low specificity and sensitivity of the less invasive methodologies that are currently available. Uncertainty about tumour diagnosis or whether a tumour has responded to treatment or has recurred can cause delays in therapeutic decisions that can impact patient outcome. Therefore, there is an urgent need to develop and validate reliable and minimally invasive biomarkers for CNS tumours that can be used alone or in combination with current clinical practices. Blood-based biomarkers can be informative in the diagnosis and monitoring of various types of cancer. However, blood-based biomarkers have proven suboptimal for analysis of CNS tumours. In contrast, circulating biomarkers in cerebrospinal fluid (CSF), including circulating tumour DNA, microRNAs and metabolites, hold promise for accurate and minimally invasive assessment of CNS tumours. This review summarises the current understanding of these three types of CSF biomarkers and their potential use in neuro-oncologic clinical practice.

Detection of the MYD88 p.L265P Mutation in the CSF of a Patient With Secondary Central Nervous System Lymphoma.

Primary Central Nervous System Lymphoma (PCNSL) and Metastatic (or Secondary) Central Nervous System Lymphoma (MCNSL) are rare central nervous system (CNS) malignancies that exhibit aggressive clinical behavior and have a poor prognosis. The majority of CNS lymphomas are histologically classified as diffuse large-B cell lymphoma (DLBCL). DLBCL harbors a high frequency of mutations in MYD88 and CD79b. The MYD88 p.L265P mutation occurs at high frequency in CNS lymphoma and is extremely rare in non-hematologic malignancies. Currently, brain biopsy is considered the gold standard for CNS lymphoma diagnosis. However, brain biopsy is invasive, carries a risk of complications, and can delay initiation of systemic therapy. Circulating tumor DNA (ctDNA) in the cerebrospinal fluid (CSF) can be utilized to detect tumor-derived mutations. Testing of CSF-ctDNA is a minimally-invasive methodology that can be used to assess the genomic alterations present in CNS malignancies. We present a case of an 82-year-old man with a history of testicular lymphoma who presented with speech difficulty and a multifocal enhancing left inferior frontal mass. Analysis for both CSF-cytology and flow cytometry did not show evidence of neoplastic cells. A brain biopsy was performed and microscopic examination showed DLBCL. We isolated CSF-ctDNA and used droplet digital PCR (ddPCR) to detect the most common lymphoma-associated mutations in MYD88, L265P, and V217F. In conjunction, we evaluated the patient-matched CNS lymphoma tissue for MYD88 mutations. We detected the MYD88 p.L265P mutation in formalin fixed paraffin embedded (FFPE) tissue from the brain biopsy and the CSF-ctDNA. In contrast, both the tumor tissue and the CSF ctDNA were negative for the MYD88 p.V217F mutation. This study shows that testing CSF ctDNA for MYD88 mutations is a potentially minimally-invasive approach to diagnosing patients with suspected CNS lymphomas.

Analysis of cerebrospinal fluid metabolites in patients with primary or metastatic central nervous system tumors.

Cancer cells have altered cellular metabolism. Mutations in genes associated with key metabolic pathways (e.g., isocitrate dehydrogenase 1 and 2, IDH1/IDH2) are important drivers of cancer, including central nervous system (CNS) tumors. Therefore, we hypothesized that the abnormal metabolic state of CNS cancer cells leads to abnormal levels of metabolites in the CSF, and different CNS cancer types are associated with specific changes in the levels of CSF metabolites. To test this hypothesis, we used mass spectrometry to analyze 129 distinct metabolites in CSF samples from patients without a history of cancer (n = 8) and with a variety of CNS tumor types (n = 23) (i.e., glioma IDH-mutant, glioma-IDH wildtype, metastatic lung cancer and metastatic breast cancer). Unsupervised hierarchical clustering analysis shows tumor-specific metabolic signatures that facilitate differentiation of tumor type from CSF analysis. We identified differences in the abundance of 43 metabolites between CSF from control patients and the CSF of patients with primary or metastatic CNS tumors. Pathway analysis revealed alterations in various metabolic pathways (e.g., glycine, choline and methionine degradation, dipthamide biosynthesis and glycolysis pathways, among others) between IDH-mutant and IDH-wildtype gliomas. Moreover, patients with IDH-mutant gliomas demonstrated higher levels of D-2-hydroxyglutarate in the CSF, in comparison to patients with other tumor types, or controls. This study demonstrates that analysis of CSF metabolites can be a clinically useful tool for diagnosing and monitoring patients with primary or metastatic CNS tumors.