Homozygous MTAP deletion in primary human glioblastoma is not associated with elevation of methylthioadenosine.

Homozygous deletion of methylthioadenosine phosphorylase (MTAP) in cancers such as glioblastoma represents a potentially targetable vulnerability. Homozygous MTAP-deleted cell lines in culture show elevation of MTAP\u27s substrate metabolite, methylthioadenosine (MTA). High levels of MTA inhibit protein arginine methyltransferase 5 (PRMT5), which sensitizes MTAP-deleted cells to PRMT5 and methionine adenosyltransferase 2A (MAT2A) inhibition. While this concept has been extensively corroborated in vitro, the clinical relevance relies on exhibiting significant MTA accumulation in human glioblastoma. In this work, using comprehensive metabolomic profiling, we show that MTA secreted by MTAP-deleted cells in vitro results in high levels of extracellular MTA. We further demonstrate that homozygous MTAP-deleted primary glioblastoma tumors do not significantly accumulate MTA in vivo due to metabolism of MTA by MTAP-expressing stroma. These findings highlight metabolic discrepancies between in vitro models and primary human tumors that must be considered when developing strategies for precision therapies targeting glioblastoma with homozygous MTAP deletion

The epigenetic evolution of glioma is determined by the IDH1 mutation status and treatment regimen

Tumor adaptation or selection is thought to underlie therapy resistance in glioma. To investigate longitudinal epigenetic evolution of gliomas in response to therapeutic pressure, we performed an epigenomic analysis of 132 matched initial and recurrent tumors from patients with IDH-wildtype (IDHwt) and IDH-mutant (IDHmut) glioma. IDHwt gliomas showed a stable epigenome over time with relatively low levels of global methylation. The epigenome of IDHmut gliomas showed initial high levels of genome-wide DNA methylation that was progressively reduced to levels similar to those of IDHwt tumors. Integration of epigenomics, gene expression, and functional genomics identified HOXD13 as a master regulator of IDHmut astrocytoma evolution. Furthermore, relapse of IDHmut tumors was accompanied by histological progression that was associated with survival, as validated in an independent cohort. Finally, the initial cell composition of the tumor microenvironment varied between IDHwt and IDHmut tumors and changed differentially following treatment, suggesting increased neo-angiogenesis and T-cell infiltration upon treatment of IDHmut gliomas. This study provides one of the largest cohorts of paired longitudinal glioma samples with epigenomic, transcriptomic, and genomic profiling and suggests that treatment of IDHmut glioma is associated with epigenomic evolution towards an IDHwt-like phenotype

The Epigenetic Evolution of Glioma Is Determined by the IDH1 Mutation Status and Treatment Regimen

Tumor adaptation or selection is thought to underlie therapy resistance in glioma. To investigate longitudinal epigenetic evolution of gliomas in response to therapeutic pressure, we performed an epigenomic analysis of 132 matched initial and recurrent tumors from patients with IDH-wildtype (IDHwt) and IDH-mutant (IDHmut) glioma. IDHwt gliomas showed a stable epigenome over time with relatively low levels of global methylation. The epigenome of IDHmut gliomas showed initial high levels of genome-wide DNA methylation that was progressively reduced to levels similar to those of IDHwt tumors. Integration of epigenomics, gene expression, and functional genomics identified HOXD13 as a master regulator of IDHmut astrocytoma evolution. Furthermore, relapse of IDHmut tumors was accompanied by histologic progression that was associated with survival, as validated in an independent cohort. Finally, the initial cell composition of the tumor microenvironment varied between IDHwt and IDHmut tumors and changed differentially following treatment, suggesting increased neoangiogenesis and T-cell infiltration upon treatment of IDHmut gliomas. This study provides one of the largest cohorts of paired longitudinal glioma samples with epigenomic, transcriptomic, and genomic profiling and suggests that treatment of IDHmut glioma is associated with epigenomic evolution toward an IDHwt-like phenotype

The Epigenetic Evolution of Glioma Is Determined by the IDH1 Mutation Status and Treatment Regimen

Tumor adaptation or selection is thought to underlie therapy resistance in glioma. To investigate longitudinal epigenetic evolution of gliomas in response to therapeutic pressure, we performed an epigenomic analysis of 132 matched initial and recurrent tumors from patients with IDH-wildtype (IDHwt) and IDH-mutant (IDHmut) glioma. IDHwt gliomas showed a stable epigenome over time with relatively low levels of global methylation. The epigenome of IDHmut gliomas showed initial high levels of genome-wide DNA methylation that was progressively reduced to levels similar to those of IDHwt tumors. Integration of epigenomics, gene expression, and functional genomics identified HOXD13 as a master regulator of IDHmut astrocytoma evolution. Furthermore, relapse of IDHmut tumors was accompanied by histologic progression that was associated with survival, as validated in an independent cohort. Finally, the initial cell composition of the tumor microenvironment varied between IDHwt and IDHmut tumors and changed differentially following treatment, suggesting increased neoangiogenesis and T-cell infiltration upon treatment of IDHmut gliomas. This study provides one of the largest cohorts of paired longitudinal glioma samples with epigenomic, transcriptomic, and genomic profiling and suggests that treatment of IDHmut glioma is associated with epigenomic evolution toward an IDHwt-like phenotype.</p

The Cyclin-like Protein Spy1 Regulates Growth and Division Characteristics of the CD133+ Population in Human Glioma

Summary The heterogeneity of brain cancers, as most solid tumors, complicates diagnosis and treatment. Identifying and targeting populations of cells driving tumorigenesis is a top priority for the cancer biology field. This is not a trivial task; considerable variance exists in the driving mutations, identifying markers, and evolutionary pressures influencing initiating cells in different individual tumors. Despite this, the ability to self-renew and differentiate must be conserved to reseed a heterogeneous tumor mass. Focusing on one example of a tumor-initiating cell population, we demonstrate that the atypical cyclin-like protein Spy1 plays a role in balancing the division properties of glioma cells with stemness properties. This mechanistic insight may provide new opportunities for therapeutic intervention of brain cancer

High-Throughput Screening of Patient-Derived Cultures Reveals Potential for Precision Medicine in Glioblastoma

Identifying drugs for the treatment of glioblastoma (GBM), a rapidly fatal disease, has been challenging. Most screening efforts have been conducted with immortalized cell lines grown with fetal bovine serum, which have little relevance to the genomic features found in GBM patients. Patient-derived neurosphere cultures, while being more physiologically relevant, are difficult to screen and therefore are only used to test a few drug candidates after initial screening efforts. Laminin has been used to generate two-dimensional cell lines from patient tumors, preserving the genomic signature and alleviating some screening hurdles. We present here the first side-by-side comparison of inhibitor sensitivity of laminin and neurosphere-grown patient-derived GBM cell lines and show that both of these culture methods result in the same pattern of inhibitor sensitivity. We used these screening methods to evaluate the dependencies of seven patient-derived cell models: three grown on laminin and four grown as neurospheres, against 56 agents in 17-point dose–response curves in 384-well format in triplicate. This allowed us to establish differential sensitivity of chemotherapeutic agents across the seven patient-derived models. We found that MEK inhibition caused patient-sample-specific growth inhibition and that bortezomib, an FDA-approved proteasome inhibitor, was potently lethal in all patient-derived models. Furthermore, the screening results led us to test the combination of the Bcl-2 inhibitor ABT-263, and the mTOR inhibitor AZD-8055, which we found to be synergistic in a subset of patient-derived GBM models. Thus, we have identified new candidate therapeutics and developed a high-throughput screening system using patient-derived GBM samples

CXCR4 increases in-vivo glioma perivascular invasion, and reduces radiation induced apoptosis: A genetic knockdown study

Glioblastoma (GBM) is a highly invasive brain tumor. Perivascular invasion, autovascularization and vascular co-option occur throughout the disease and lead to tumor invasion and progression. The molecular basis for perivascular invasion, i.e., the interaction of glioma tumor cells with endothelial cells is not well characterized. Recent studies indicate that glioma cells have increased expression of CXCR4. We investigated the in-vivo role of CXCR4 in perivascular invasion of glioma cells using shRNA-mediated knock down of CXCR4. We show that primary cultures of human glioma stem cells HF2303 and mouse glioma GL26-Cit cells exhibit significant migration towards human (HBMVE) and mouse (MBVE) brain microvascular endothelial cells. Blocking CXCR4 on tumor cells with AMD3100 in-vitro, inhibits migration of GL26-Cit and HF2303 toward MBVE and HBMVE cells. Additionally, genetic down regulation of CXCR4 in mouse glioma GL26-Cit cells inhibits their in-vitro migration towards MBVE cells; in an in-vivo intracranial mouse model, these cells display reduced tumor growth and perivascular invasion, leading to increased survival. Quantitative analysis of brain sections showed that CXCR4 knockdown tumors are less invasive. Lastly, we tested the effects of radiation on CXCR4 knock down GL26-Cit cells in an orthotopic brain tumor model. Radiation treatment increased apoptosis of CXCR4 downregulated tumor cells and prolonged median survival. In summary, our data suggest that CXCR4 signaling is critical for perivascular invasion of GBM cells and targeting this receptor makes tumors less invasive and more sensitive to radiation therapy. Combination of CXCR4 knock down and radiation treatment might improve the efficacy of GBM therapy

A novel natural product compound enhances cAMP-regulated chloride conductance of cells expressing CFTRΔf508

Background: Cystic fibrosis (CF) results from mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which encodes a chloride channel localized at the plasma membrane of diverse epithelia. The most common mutation leading to CF, ΔF508, occurs in the first nucleotide-binding domain (NBD1) of CFTR. The ΔF50