Oncolytic virotherapy is a treatment modality that uses native or genetically modified viruses that selectively replicate in and kill tumor cells. Viruses represent a type of pathogen-associated molecular pattern and thereby induce the up-regulation of dozens of cytokines via activating the host innate immune system. Second mitochondria-derived activator of caspases (Smac) mimetic compounds (SMCs), which antagonize the function of inhibitor of apoptosis proteins (IAPs) and induce apoptosis, sensitize tumor cells to multiple cytokines. Therefore, we sought to determine whether SMCs sensitize tumor cells to cytokines induced by the oncolytic M1 virus, thus enhancing a bystander killing effect. Here, we report that SMCs potentiate the oncolytic effect of M1 in vitro, in vivo, and ex vivo. This strengthened oncolytic efficacy resulted from the enhanced bystander killing effect caused by the M1 virus via cytokine induction. Through a microarray analysis and subsequent validation using recombinant cytokines, we identified IL-8, IL-1A, and TRAIL as the key cytokines in the bystander killing effect. Furthermore, SMCs increased the replication of M1, and the accumulation of virus protein induced irreversible endoplasmic reticulum stress- and c-Jun N-terminal kinase-mediated apoptosis. Nevertheless, the combined treatment with M1 and SMCs had little effect on normal and human primary cells. Because SMCs selectively and significantly enhance the bystander killing effect and the replication of oncolytic virus M1 specifically in cancer cells, this combined treatment may represent a promising therapeutic strategy.

The RB1 gene is the first described human tumor suppressor gene and plays an integral role in the development of retinoblastoma, a pediatric malignancy of the eye. Since its discovery, the stepwise characterization and cloning of RB1 have laid the foundation for numerous advances in the understanding of tumor suppressor genes, retinoblastoma tumorigenesis, and inheritance. Knowledge of RB1 led to a paradigm shift in the field of cancer genetics, including widespread acceptance of the concept of tumor suppressor genes, and has provided crucial diagnostic and prognostic information through genetic testing for patients affected by retinoblastoma. This article reviews the long history of RB1 gene research, characterization, and cloning, and also discusses recent advances in retinoblastoma genetics that have grown out of this foundational work.

Glioblastoma multiforme (GBM) is an aggressive cancer without currently effective therapies. Radiation and temozolomide (radio/TMZ) resistance are major contributors to cancer recurrence and failed GBM therapy. Heat shock proteins (HSPs), through regulation of extracellular matrix (ECM) remodeling and epithelial mesenchymal transition (EMT), provide mechanistic pathways contributing to the development of GBM and radio/TMZ-resistant GBM. The Friend leukemia integration 1 (Fli-1) signaling network has been implicated in oncogenesis in GBM, making it an appealing target for advancing novel therapeutics. Fli-1 is linked to oncogenic transformation with up-regulation in radio/TMZ-resistant GBM, transcriptionally regulating HSPB1. This link led us to search for targeted molecules that inhibit Fli-1. Expression screening for Fli-1 inhibitors identified lumefantrine, an antimalarial drug, as a probable Fli-1 inhibitor. Docking and isothermal calorimetry titration confirmed interaction between lumefantrine and Fli-1. Lumefantrine promoted growth suppression and apoptosis in vitro in parental and radio/TMZ-resistant GBM and inhibited tumor growth without toxicity in vivo in U87MG GBM and radio/TMZ-resistant GBM orthotopic tumor models. These data reveal that lumefantrine, an FDA-approved drug, represents a potential GBM therapeutic that functions through inhibition of the Fli-1/HSPB1/EMT/ECM remodeling protein networks.

Cancer cells adapt their signaling in response to nutrient availability. To uncover the mechanisms regulating this process and its functional consequences, we interrogated cell lines, mouse tumor models, and clinical samples of glioblastoma (GBM), the highly lethal brain cancer. We discovered that glucose or acetate is required for epidermal growth factor receptor vIII (EGFRvIII), the most common growth factor receptor mutation in GBM, to activate mechanistic target of rapamycin complex 2 (mTORC2) and promote tumor growth. Glucose or acetate promoted growth factor receptor signaling through acetyl-CoA-dependent acetylation of Rictor, a core component of the mTORC2 signaling complex. Remarkably, in the presence of elevated glucose levels, Rictor acetylation is maintained to form an autoactivation loop of mTORC2 even when the upstream components of the growth factor receptor signaling pathway are no longer active, thus rendering GBMs resistant to EGFR-, PI3K (phosphoinositide 3-kinase)-, or AKT (v-akt murine thymoma viral oncogene homolog)-targeted therapies. These results demonstrate that elevated nutrient levels can drive resistance to targeted cancer treatments and nominate mTORC2 as a central node for integrating growth factor signaling with nutrient availability in GBM.

Melanoma differentiation-associated gene-7/interleukin-24 (mda-7/IL-24) is a multifunctional cytokine displaying broad-spectrum anticancer activity in vitro or in vivo in preclinical animal cancer models and in a phase 1/2 clinical trial in patients with advanced cancers. mda-7/IL-24 targets specific miRNAs, including miR-221 and miR-320, for down-regulation in a cancer-selective manner. We demonstrate that mda-7/IL-24, administered through a replication incompetent type 5 adenovirus (Ad.mda-7) or with His-MDA-7/IL-24 protein, down-regulates DICER, a critical regulator in miRNA processing. This effect is specific for mature miR-221, as it does not affect Pri-miR-221 expression, and the DICER protein, as no changes occur in other miRNA processing cofactors, including DROSHA, PASHA, or Argonaute. DICER is unchanged by Ad.mda-7/IL-24 in normal immortal prostate cells, whereas Ad.mda-7 down-regulates DICER in multiple cancer cells including glioblastoma multiforme and prostate, breast, lung, and liver carcinoma cells. MDA-7/IL-24 protein down-regulates DICER expression through canonical IL-20/IL-22 receptors. Gain- and loss-of-function studies confirm that overexpression of DICER rescues deregulation of miRNAs by mda-7/IL-24, partially rescuing cancer cells from mda-7/IL-24-mediated cell death. Stable overexpression of DICER in cancer cells impedes Ad.mda-7 or His-MDA-7/IL-24 inhibition of cell growth, colony formation, PARP cleavage, and apoptosis. In addition, stable overexpression of DICER renders cancer cells more resistant to Ad.mda-7 inhibition of primary and secondary tumor growth. MDA-7/IL-24-mediated regulation of DICER is reactive oxygen species-dependent and mediated by melanogenesis-associated transcription factor. Our research uncovers a distinct role of mda-7/IL-24 in the regulation of miRNA biogenesis through alteration of the MITF-DICER pathway.

In cancer, aberrant growth factor receptor signaling reprograms cellular metabolism and global gene transcription to drive aggressive growth, but the underlying mechanisms are not well-understood. Here we show that in the highly lethal brain tumor glioblastoma (GBM), mTOR complex 2 (mTORC2), a critical core component of the growth factor signaling system, couples acetyl-CoA production with nuclear translocation of histone-modifying enzymes including pyruvate dehydrogenase and class IIa histone deacetylases to globally alter histone acetylation. Integrated analyses in orthotopic mouse models and in clinical GBM samples reveal that mTORC2 controls iron metabolisms via histone H3 acetylation of the iron-related gene promoter, promoting tumor cell survival. These results nominate mTORC2 as a critical epigenetic regulator of iron metabolism in cancer.

EGFR is the most common genetically altered oncogene in glioblastoma (GBM), but small-molecule EGFR tyrosine kinase inhibitors (TKI) have failed to yield durable clinical benefit. Here, we show that in two novel model systems of acquired resistance to EGFR TKIs, elevated expression of urokinase plasminogen activator (uPA) drives signaling through the MAPK pathway, which results in suppression of the proapoptotic BCL2-family member protein BIM (BCL2L11). In patient-derived GBM cells and genetic GBM models, uPA is shown to suppress BIM levels through ERK1/2 phosphorylation, which can be reversed by siRNA-mediated knockdown of uPA. TKI-resistant GBMs are resensitized to EGFR TKIs by pharmacologic inhibition of MEK or a BH3 mimetic drug to replace BIM function. A link between the uPA-uPAR-ERK1/2 pathway and BIM has not been previously demonstrated in GBM, and involvement of this signaling axis in resistance provides rationale for a new strategy to target EGFR TKI-resistant GBM.

Somatic copy number alterations (SCNAs), generally (1) losses containing interferons and interferon-pathway genes, many on chromosome 9p, predict immune-cold, immune checkpoint therapy (ICT)-resistant tumors (2); however, genomic regions mediating these effects are unclear and probably tissue specific. Previously, 9p21.3 loss was found to be an early genetic driver of human papillomavirus-negative (HPV^-) head and neck squamous cancer (HNSC), associated with an immune-cold tumor microenvironment (TME) signal, and recent evidence suggested that this TME-cold phenotype was greatly enhanced with 9p21 deletion size, notably encompassing band 9p24.1 (3). Here, we report multi-omic, -threshold and continuous-variable dissection of 9p21 and 9p24 loci (including depth and degree of somatic alteration of each band at each locus, and each gene at each band) and TME of four HPV^- HNSC cohorts. Preferential 9p24 deletion, CD8 T-cell immune-cold associations were observed, driven by 9p24.1 loss, and in turn by an essential telomeric regulatory gene element, JAK2-CD274. Surprisingly, same genetic region gains were immune hot. Related 9p21-TME analyses were less evident. Inherent 9p-band-level influences on anti-PD1 ICT survival rates, coincident with TME patterns, were also observed. At a 9p24.1 whole-transcriptome expression threshold of 60th percentile, ICT survival rate exceeded that of lower expression percentiles and of chemotherapy; below this transcript threshold, ICT survival was inferior to chemotherapy, the latter unaffected by 9p24.1 expression level (P-values < 0.01, including in a PD-L1 immunohistochemistry-positive patient subgroup). Whole-exome analyses of 10 solid-tumor types suggest that these 9p-related ICT findings could be relevant to squamous cancers, in which 9p24.1 gain/immune-hot associations exist.

Glioblastoma multiforme (GBM) is an intractable tumor despite therapeutic advances, principally because of its invasive properties. Radiation is a staple in therapeutic regimens, although cells surviving radiation can become more aggressive and invasive. Subtraction hybridization identified melanoma differentiation-associated gene 9 [MDA-9/Syntenin; syndecan-binding protein (SDCBP)] as a differentially regulated gene associated with aggressive cancer phenotypes in melanoma. MDA-9/Syntenin, a highly conserved double-PDZ domain-containing scaffolding protein, is robustly expressed in human-derived GBM cell lines and patient samples, with expression increasing with tumor grade and correlating with shorter survival times and poorer response to radiotherapy. Knockdown of MDA-9/Syntenin sensitizes GBM cells to radiation, reducing postradiation invasion gains. Radiation induces Src and EGFRvIII signaling, which is abrogated through MDA-9/Syntenin down-regulation. A specific inhibitor of MDA-9/Syntenin activity, PDZ1i (113B7), identified through NMR-guided fragment-based drug design, inhibited MDA-9/Syntenin binding to EGFRvIII, which increased following radiation. Both genetic (shmda-9) and pharmacological (PDZ1i) targeting of MDA-9/Syntenin reduced invasion gains in GBM cells following radiation. Although not affecting normal astrocyte survival when combined with radiation, PDZ1i radiosensitized GBM cells. PDZ1i inhibited crucial GBM signaling involving FAK and mutant EGFR, EGFRvIII, and abrogated gains in secreted proteases, MMP-2 and MMP-9, following radiation. In an in vivo glioma model, PDZ1i resulted in smaller, less invasive tumors and enhanced survival. When combined with radiation, survival gains exceeded radiotherapy alone. MDA-9/Syntenin (SDCBP) provides a direct target for therapy of aggressive cancers such as GBM, and defined small-molecule inhibitors such as PDZ1i hold promise to advance targeted brain cancer therapy.

EGFR is the most common genetically altered oncogene in glioblastoma (GBM), but small-molecule EGFR tyrosine kinase inhibitors (TKI) have failed to yield durable clinical benefit. Here, we show that in two novel model systems of acquired resistance to EGFR TKIs, elevated expression of urokinase plasminogen activator (uPA) drives signaling through the MAPK pathway, which results in suppression of the proapoptotic BCL2-family member protein BIM (BCL2L11). In patient-derived GBM cells and genetic GBM models, uPA is shown to suppress BIM levels through ERK1/2 phosphorylation, which can be reversed by siRNA-mediated knockdown of uPA. TKI-resistant GBMs are resensitized to EGFR TKIs by pharmacologic inhibition of MEK or a BH3 mimetic drug to replace BIM function. A link between the uPA-uPAR-ERK1/2 pathway and BIM has not been previously demonstrated in GBM, and involvement of this signaling axis in resistance provides rationale for a new strategy to target EGFR TKI-resistant GBM.