Anh Nhat Tran

NOS Expression and NO Function in Glioma and Implications for Patient Therapies.

SIGNIFICANCEnGliomas are central nervous system tumors that primarily occur in the brain and arise from glial cells. Gliomas include the most common malignant brain tumor in adults known as grade IV astrocytoma, or glioblastoma (GBM). GBM is a deadly disease for which the most significant advances in treatment offer an improvement in survival of only ∼2 months.nnnCRITICAL ISSUESnTo develop novel treatments and improve patient outcomes, we and others have sought to determine the role of molecular signals in gliomas. Recent Advances: One signaling molecule that mediates important biologies in glioma is the free radical nitric oxide (NO). In glioma cells and the tumor microenvironment, NO is produced by three isoforms of nitric oxide synthase (NOS), NOS1, NOS2, and NOS3. NO and NOS affect glioma growth, invasion, angiogenesis, immunosuppression, differentiation state, and therapeutic resistance.nnnFUTURE DIRECTIONSnThese multifaceted effects of NO and NOS on gliomas both in vitro and in vivo suggest the potential of modulating the pathway for antiglioma patient therapies. Antioxid. Redox Signal. 26, 986-999.

Addition of carbonic anhydrase 9 inhibitor SLC-0111 to temozolomide treatment delays glioblastoma growth in vivo

Tumor microenvironments can promote stem cell maintenance, tumor growth, and therapeutic resistance, findings linked by the tumor-initiating cell hypothesis. Standard of care for glioblastoma (GBM) includes temozolomide chemotherapy, which is not curative, due, in part, to residual therapy-resistant brain tumor-initiating cells (BTICs). Temozolomide efficacy may be increased by targeting carbonic anhydrase 9 (CA9), a hypoxia-responsive gene important for maintaining the altered pH gradient of tumor cells. Using patient-derived GBM xenograft cells, we explored whether CA9 and CA12 inhibitor SLC-0111 could decrease GBM growth in combination with temozolomide or influence percentages of BTICs after chemotherapy. In multiple GBMs, SLC-0111 used concurrently with temozolomide reduced cell growth and induced cell cycle arrest via DNA damage in vitro. In addition, this treatment shifted tumor metabolism to a suppressed bioenergetic state in vivo. SLC-0111 also inhibited the enrichment of BTICs after temozolomide treatment determined via CD133 expression and neurosphere formation capacity. GBM xenografts treated with SLC-0111 in combination with temozolomide regressed significantly, and this effect was greater than that of temozolomide or SLC-0111 alone. We determined that SLC-0111 improves the efficacy of temozolomide to extend survival of GBM-bearing mice and should be explored as a treatment strategy in combination with current standard of care.

The pro-tumorigenic effects of metabolic alterations in glioblastoma including brain tumor initiating cells

De-regulated cellular energetics is an emerging hallmark of cancer with alterations to glycolysis, oxidative phosphorylation, the pentose phosphate pathway, lipid oxidation and synthesis and amino acid metabolism. Understanding and targeting of metabolic reprogramming in cancers may yield new treatment options, but metabolic heterogeneity and plasticity complicate this strategy. One highly heterogeneous cancer for which current treatments ultimately fail is the deadly brain tumor glioblastoma. Therapeutic resistance, within glioblastoma and other solid tumors, is thought to be linked to subsets of tumor initiating cells, also known as cancer stem cells. Recent profiling of glioblastoma and brain tumor initiating cells reveals changes in metabolism, as compiled here, that may be more broadly applicable. We will summarize the profound role for metabolism in tumor progression and therapeutic resistance and discuss current approaches to target glioma metabolism to improve standard of care.

Modeling Physiologic Microenvironments in Three-Dimensional Microtumors Maintains Brain Tumor Initiating Cells

Development of effective novel anti-tumor treatments will require improved in vitro models that incorporate physiologic microenvironments and maintain intratumoral heterogeneity, including tumor initiating cells. Brain tumor initiating cells (BTIC) are a target for cancer therapy, because BTICs are highly tumorigenic and contribute to tumor angiogenesis, invasion, and therapeutic resistance. Current leading studies rely on BTIC isolation from patient-derived xenografts followed by propagation as neurospheres. As this process is expensive and time-consuming, we determined whether three-dimensional microtumors were an alternative in vitro method for modeling tumor growth via BITC maintenance and/or enrichment. Brain tumor cells were grown as neurospheres or as microtumors produced using the human-derived biomatrix HuBiogel™ and maintained with physiologically relevant microenvironments. BITC percentages were determined using cell surface marker expression, label retention, and neurosphere formation capacity. Our data demonstrate that expansion of brain tumor cells as hypoxic and nutrient-restricted microtumors significantly increased the percentage of both CD133+ and CFSEhigh cells. We further demonstrate that BTIC-marker positive cells isolated from microtumors maintained neurosphere formation capacity in the in vitro limiting dilution assay and tumorigenic potential in vivo. These data demonstrate that microtumors can be a useful three-dimensional biological model for the study of BTIC maintenance and targeting.

HPAanalyze: An R Package that Facilitate the Retrieval and Analysis of The Human Protein Atlas Data

Background The Human Protein Atlas aims to map human proteins via multiple technologies including imaging, proteomics and transcriptomics. Access of the HPA data is mainly via web-based interface allowing view of individual proteins, which may not be optimal for data analysis of a gene set, or automatic retrieval of original images. Results HPAanalyze is an R package for retrieving and performing exploratory data analysis from HPA. It provides functionality for importing data tables and xml files from HPA, exporting and visualizing data, as well as download all staining images of interest. The package is free, open source, and available via GitHub. Conclusions HPAanalyze integrates into the R workflow via the tidyverse philosophy and data structures, and can be used in combination with Bioconductor packages for easy analysis of HPA data.

Reactive species balance via GTP cyclohydrolase I regulates glioblastoma growth and tumor initiating cell maintenance

BackgroundnDepending on the level, differentiation state, and tumor stage, reactive nitrogen and oxygen species inhibit or increase cancer growth and tumor initiating cell maintenance. The rate-limiting enzyme in a pathway that can regulate reactive species production but has not been thoroughly investigated in glioblastoma (GBM; grade IV astrocytoma) is guanosine triphosphate (GTP) cyclohydrolase 1 (GCH1). We sought to define the role of GCH1 in the regulation of GBM growth and brain tumor initiating cell (BTIC) maintenance.nnnMethodsnWe examined GCH1 mRNA and protein expression in patient-derived xenografts, clinical samples, and glioma gene expression datasets. GCH1 levels were modulated using lentiviral expression systems, and effects on cell growth, self-renewal, reactive species production, and survival in orthotopic patient-derived xenograft models were determined.nnnResultsnGCH1 was expressed in GBMs with elevated but not exclusive RNA and protein levels in BTICs in comparison to non-BTICs. Overexpression of GCH1 in GBM cells increased cell growth in vitro and decreased survival in an intracranial GBM mouse model. In converse experiments, GCH1 knockdown with short hairpin RNA led to GBM cell growth inhibition and reduced self-renewal in association with decreased CD44 expression. GCH1 was critical for controlling reactive species balance, including suppressing reactive oxygen species production, which mediated GCH1 cell growth effects. In silico analyses demonstrated that higher GCH1 levels in glioma patients correlate with higher glioma grade, recurrence, and worse survival.nnnConclusionsnGCH1 expression in established GBMs is pro-tumorigenic, causing increased growth due, in part, to promotion of BTIC maintenance and suppression of reactive oxygen species.

Identification of Compounds That Decrease Glioblastoma Growth and Glucose Uptake in Vitro

Tumor heterogeneity has hampered the development of novel effective therapeutic options for aggressive cancers, including the deadly primary adult brain tumor glioblastoma (GBM). Intratumoral heterogeneity is partially attributed to the tumor initiating cell (TIC) subset that contains highly tumorigenic, stem-like cells. TICs display metabolic plasticity but can have a reliance on aerobic glycolysis. Elevated expression of GLUT1 and GLUT3 is present in many cancer types, with GLUT3 being preferentially expressed in brain TICs (BTICs) to increase survival in low nutrient tumor microenvironments, leading to tumor maintenance. Through structure-based virtual screening (SBVS), we identified potential novel GLUT inhibitors. The screening of 13 compounds identified two that preferentially inhibit the growth of GBM cells with minimal toxicity to non-neoplastic astrocytes and neurons. These compounds, SRI-37683 and SRI-37684, also inhibit glucose uptake and decrease the glycolytic capacity and glycolytic reserve capacity of GBM patient-derived xenograft (PDX) cells in glycolytic stress test assays. Our results suggest a potential new therapeutic avenue to target metabolic reprogramming for the treatment of GBM, as well as other tumor types, and the identified novel inhibitors provide an excellent starting point for further lead development.