Jason K. Sa

Transglutaminase 2 Inhibition Reverses Mesenchymal Transdifferentiation of Glioma Stem Cells by Regulating C/EBPβ Signaling

Necrosis is a hallmark of glioblastoma (GBM) and is responsible for poor prognosis and resistance to conventional therapies. However, the molecular mechanisms underlying necrotic microenvironment-induced malignancy of GBM have not been elucidated. Here, we report that transglutaminase 2 (TGM2) is upregulated in the perinecrotic region of GBM and triggered mesenchymal (MES) transdifferentiation of glioma stem cells (GSC) by regulating master transcription factors (TF), such as C/EBPβ, TAZ, and STAT3. TGM2 expression was induced by macrophages/microglia-derived cytokines via NF-κB activation and further degraded DNA damage-inducible transcript 3 (GADD153) to induce C/EBPβ expression, resulting in expression of the MES transcriptome. Downregulation of TGM2 decreased sphere-forming ability, tumor size, and radioresistance and survival in a xenograft mouse model through a loss of the MES signature. A TGM2-specific inhibitor GK921 blocked MES transdifferentiation and showed significant therapeutic efficacy in mouse models of GSC. Moreover, TGM2 expression was significantly increased in recurrent MES patients and inversely correlated with patient prognosis. Collectively, our results indicate that TGM2 is a key molecular switch of necrosis-induced MES transdifferentiation and an important therapeutic target for MES GBM. Cancer Res; 77(18); 4973-84. ©2017 AACR.

Pharmacogenomic landscape of patient-derived tumor cells informs precision oncology therapy

Outcomes of anticancer therapy vary dramatically among patients due to diverse genetic and molecular backgrounds, highlighting extensive intertumoral heterogeneity. The fundamental tenet of precision oncology defines molecular characterization of tumors to guide optimal patient-tailored therapy. Towards this goal, we have established a compilation of pharmacological landscapes of 462 patient-derived tumor cells (PDCs) across 14 cancer types, together with genomic and transcriptomic profiling in 385 of these tumors. Compared with the traditional long-term cultured cancer cell line models, PDCs recapitulate the molecular properties and biology of the diseases more precisely. Here, we provide insights into dynamic pharmacogenomic associations, including molecular determinants that elicit therapeutic resistance to EGFR inhibitors, and the potential repurposing of ibrutinib (currently used in hematological malignancies) for EGFR-specific therapy in gliomas. Lastly, we present a potential implementation of PDC-derived drug sensitivities for the prediction of clinical response to targeted therapeutics using retrospective clinical studies.Analysis of genomic and transcriptomic data from 462 patient-derived tumor cell (PDC) samples across 14 cancer types, along with pharmacological responses to 60 agents, indicates that PDC-derived drug sensitivities might be predictive of clinical response to targeted therapies.

Glioma through the looking GLASS: molecular evolution of diffuse gliomas and the Glioma Longitudinal Analysis Consortium

Abstract Adult diffuse gliomas are a diverse group of brain neoplasms that inflict a high emotional toll on patients and their families. The Cancer Genome Atlas and similar projects have provided a comprehensive understanding of the somatic alterations and molecular subtypes of glioma at diagnosis. However, gliomas undergo significant cellular and molecular evolution during disease progression. We review the current knowledge on the genomic and epigenetic abnormalities in primary tumors and after disease recurrence, highlight the gaps in the literature, and elaborate on the need for a new multi-institutional effort to bridge these knowledge gaps and how the Glioma Longitudinal Analysis Consortium (GLASS) aims to systemically catalog the longitudinal changes in gliomas. The GLASS initiative will provide essential insights into the evolution of glioma toward a lethal phenotype, with the potential to reveal targetable vulnerabilities and, ultimately, improved outcomes for a patient population in need.

Identification of genomic and molecular traits that present therapeutic vulnerability to HGF-targeted therapy in glioblastoma

BACKGROUND Cancer is a complex disease with profound genomic alterations and extensive heterogeneity. Recent studies on large-scale genomics have shed light on the impact of core oncogenic pathways, which are frequently dysregulated in a wide spectrum of cancer types. Aberrant activation of the hepatocyte growth factor (HGF) signaling axis has been associated with promoting various oncogenic programs during tumor initiation, progression, and treatment resistance. As a result, HGF-targeted therapy has emerged as an attractive therapeutic approach. However, recent clinical trials involving HGF-targeted therapies have demonstrated rather disappointing results. Thus, an alternative, in-depth assessment of new patient stratification is necessary to shift the current clinical course. METHODS To address such challenges, we have evaluated the therapeutic efficacy of YYB-101, an HGF-neutralizing antibody, in a series of primary glioblastoma stem cells (GSCs) both in vitro and in vivo. Furthermore, we performed genome and transcriptome analysis to determine genetic and molecular traits that exhibit therapeutic susceptibility to HGF-mediated therapy. RESULTS We have identified several differentially expressed genes, including MET, KDR, and SOX3, which are associated with tumor invasiveness, malignancy, and unfavorable prognosis in glioblastoma patients. We also demonstrated the HGF-MET signaling axis as a key molecular determinant in GSC invasion, and we discovered that a significant association in HGF expression existed between mesenchymal phenotype and immune cell recruitment. CONCLUSIONS Upregulation of MET and mesenchymal cellular state are essential in generating HGF-mediated therapeutic responses. Our results provide an important framework for evaluating HGF-targeted therapy in future clinical settings.

A tension-mediated glycocalyx–integrin feedback loop promotes mesenchymal-like glioblastoma

Glioblastoma multiforme (GBMs) are recurrent lethal brain tumours. Recurrent GBMs often exhibit mesenchymal, stem-like phenotypes that could explain their resistance to therapy. Analyses revealed that recurrent GBMs have increased tension and express high levels of glycoproteins that increase the bulkiness of the glycocalyx. Studies showed that a bulky glycocalyx potentiates integrin mechanosignalling and tissue tension and promotes a mesenchymal, stem-like phenotype in GBMs. Gain- and loss-of-function studies implicated integrin mechanosignalling as an inducer of GBM growth, survival, invasion and treatment resistance, and a mesenchymal, stem-like phenotype. Mesenchymal-like GBMs were highly contractile and expressed elevated levels of glycoproteins that expanded their glycocalyx, and they were surrounded by a stiff extracellular matrix that potentiated integrin mechanosignalling. Our findings suggest that there is a dynamic and reciprocal link between integrin mechanosignalling and a bulky glycocalyx, implying a causal link towards a mesenchymal, stem-like phenotype in GBMs. Strategies to ameliorate GBM tissue tension offer a therapeutic approach to reduce mortality due to GBM.Barnes et al. report a dynamic and reciprocal crosstalk between tissue tension and glycocalyx bulkiness that promotes a mesenchymal, stem-like phenotype in GBM.

Comparison of 1p and 19q status of glioblastoma by whole exome sequencing, array-comparative genomic hybridization, and fluorescence in situ hybridization

According to the 2016 World Health Organization classification of tumors of the central nervous system, detecting 1p/19q co-deletion became essential in clinical neuropathology for gliomas with oligodendroglioma-like morphology. Here, we assessed genomic profiles of glioblastoma in 80 cases including 1p/19q status using fluorescent in situ hybridization (FISH), array-comparative genomic hybridization (aCGH), and/or whole exome sequencing (WES). Paraffin-embedded tumor tissues were subjected to FISH analysis, and the corresponding frozen tissues from the same tumors were evaluated for aCGH and/or WES for 1p/19q co-deletion and other genetic parameters, which included IDH1-R132H, ATRX, TP53, CIC, and NOTCH1 mutations and MGMT methylation status. We also evaluated correlations between 1p/19q co-deletion status and molecular markers or clinical outcomes. The FISH analyses revealed 1p/19q co-deletion in two cases, isolated deletion of 1p in six cases, and 19q in two cases, whereas the aCGH and WES results showed isolated deletion of 19q in four cases and 19 monosomy in only one case. Eleven cases showed discordant 1p/19q results between aCGH/WES and FISH analysis, and in most of them, 1p and/or 19q deletion on FISH analysis corresponded to the partial deletions at 1p36 and/or 19q13 on aCGH/WES. Our cohort exhibited IDH1-R132H mutations (5.4%), MGMT promotor methylation (34.6%), and mutations in ATRX (9.5%), TP53 (33.3%), and NOTCH1 (3.8%) but not in CIC (0%). In addition, MGMT methylation and ATRX mutation were significantly associated with clinical prognosis. In glioblastomas, partial deletions of 1p36 and/or 19q13 were uncommon, some of which appeared as 1p and/or 19q deletions on FISH analysis.

Identification of transcriptome signature for predicting clinical response to bevacizumab in recurrent glioblastoma

Glioblastomas are among the most fatal brain tumors. Although no effective treatment option is available for recurrent glioblastomas (GBMs), a subset of patients evidently derived clinical benefit from bevacizumab, a monoclonal antibody against vascular endothelial growth factor. We retrospectively reviewed patients with recurrent GBM who received bevacizumab to identify biomarkers for predicting clinical response to bevacizumab. Following defined criteria, the patients were categorized into two clinical response groups, and their genetic and transcriptomic results were compared. Angiogenesis‐related gene sets were upregulated in both responders and nonresponders, whereas genes for each corresponding angiogenesis pathway were distinct from one another. Two gene sets were made, namely, the nonresponder angiogenesis gene set (NAG) and responder angiogenesis gene set (RAG), and then implemented in independent GBM cohort to validate our dataset. A similar association between the corresponding gene set and survival was observed. In NAG, COL4A2 was associated with a poor clinical outcome in bevacizumab‐treated patients. This study demonstrates that angiogenesis‐associated gene sets are composed of distinct subsets with diverse biological roles and they represent different clinical responses to anti‐angiogenic therapy. Enrichment of a distinct angiogenesis pathway may serve as a biomarker to predict patients who will derive a clinical benefit from bevacizumab.

Abstract 4664: Single-cell transcriptome analysis on lung squamous cell carcinoma derived from a patient

Distinct genomic signature of a patient9s cancer could provide a clue to design and predict accurate therapeutic responses to targeted medicine. Current approaches regarding heterogeneous cancer cells en masse as a pooled population, however, hardly reflect complete genomic landscape of tumor diversity, missing potentially important minor clones9 implications regarding metastasis and drug resistance. Here we report a comprehensive analysis to discover different patterns of subclonal behaviors in functional modalities and signaling pathways out of heterogeneous population derived from a patient with lung squamous cell carcinoma, using single-cell mRNA sequencing. As a result of nonnegative matrix factorization as a clustering, we identified discrete 4 subgroups with a major group in accordance with overall expression profile of a pooled population of cells, and the other 3 minor groups that were masked in a pooled sample. Genomic variants were also reliably identified and compared across single cells. We integrated these genomic signatures from single cells to address the different responses in drug screening. Together, our approach provides insights into the more accurate strategy to identify minor but potentially important subclones that are relevant to drug resistance. Citation Format: Kyu-Tae Kim, Hye-Won Lee, Kyeung Min Joo, Sang-Chul Kim, Jin-Ku Lee, Nakho Chang, Jason K. Sa, Yu Jin Cho, Do-Hyun Nam, Hae-Ock Lee, Woong-Yang Park. Single-cell transcriptome analysis on lung squamous cell carcinoma derived from a patient. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4664. doi:10.1158/1538-7445.AM2014-4664