Highlights d Rapid establishment of glioblastoma organoids (GBOs) in a defined medium with biobank d GBOs maintain parental tumor cellular heterogeneity, gene expression, and mutations d GBO transplantation exhibits efficient engraftment and aggressive infiltration d Tumor-specific treatment responses in GBOs to drugs and CART cells

Gold nanoparticles (AuNPs) have generated interest as both imaging and therapeutic agents. AuNPs are attractive for imaging applications since they are nontoxic and provide nearly three times greater X-ray attenuation per unit weight than iodine. As therapeutic agents, AuNPs can sensitize tumor cells to ionizing radiation. To create a nanoplatform that could simultaneously exhibit long circulation times, achieve appreciable tumor accumulation, generate computed tomography (CT) image contrast, and serve as a radiosensitizer, gold-loaded polymeric micelles (GPMs) were prepared. Specifically, 1.9 nm AuNPs were encapsulated within the hydrophobic core of micelles formed with the amphiphilic diblock copolymer poly(ethylene glycol)-b-poly(ε-capralactone). GPMs were produced with low polydispersity and mean hydrodynamic diameters ranging from 25 to 150 nm. Following intravenous injection, GPMs provided blood pool contrast for up to 24 h and improved the delineation of tumor margins via CT. Thus, GPM-enhanced CT imaging was used to guide radiation therapy delivered via a small animal radiation research platform. In combination with the radiosensitizing capabilities of gold, tumor-bearing mice exhibited a 1.7-fold improvement in the median survival time, compared with mice receiving radiation alone. It is envisioned that translation of these capabilities to human cancer patients could guide and enhance the efficacy of radiation therapy.

Blood tests to detect circulating tumor cells (CTC) offer great potential to monitor disease status, gauge prognosis, and guide treatment decisions for patients with cancer. For patients with brain tumors, such as aggressive glioblastoma multiforme, CTC assays are needed that do not rely on expression of cancer cell surface biomarkers like epithelial cell adhesion molecules that brain tumors tend to lack. Here, we describe a strategy to detect CTC based on telomerase activity, which is elevated in nearly all tumor cells but not normal cells. This strategy uses an adenoviral detection system that is shown to successfully detect CTC in patients with brain tumors. Clinical data suggest that this assay might assist interpretation of treatment response in patients receiving radiotherapy, for example, to differentiate pseudoprogression from true tumor progression. These results support further development of this assay as a generalized method to detect CTC in patients with cancer.

Grb2-associated binder-1 (Gab1) is a multisite docking protein containing a pleckstrin homology (PH) domain, multiple potential tyrosine phosphorylation sites, and several proline-rich sequences. Gab1 becomes tyrosinephosphorylated in cells stimulated with growth factors, cytokines, and ligands for G protein-coupled receptors. A major Gab1-binding protein detected in cells treated with extracellular stimuli is the tyrosine phosphatase, SHP2. Although the role of SHP2-Gab1 interaction in cell signaling has not yet been characterized, SHP2 is known to mediate mitogen-activated protein (MAP) kinase activation induced by the epidermal growth factor (EGF). However, the mechanism by which the SHP2 phosphatase exerts a positive signaling role remains obscure. In this study, we prepared Gab1 mutants lacking the SHP2 binding site (Gab1Y627F), the phosphatidylinositol 3-kinase (PI3K) binding sites (Gab1⌬PI3K), and the PH domain (Gab1⌬PH). Expression of Gab1Y627F blocked the extracellular signal-regulated kinase-2 (ERK2) activation by lysophosphatidic acid (LPA) and EGF. Conversely, expression of the wild-type Gab1 in HEK293 cells augmented the LPA receptor Edg2-mediated ERK2 activation. Whereas the PH domain was required for Gab1 mediation of ERK2 activation by LPA, it was not essential for EGF-induced ERK2 activation. Expression of Gab1⌬PI3K had no apparent effect on ERK2 activation by LPA and EGF in the cells that we have examined. These results establish a role for Gab1 in the LPA-induced MAP kinase pathway and clearly demonstrate that Gab1-SHP2 interaction is essential for ERK2 activation by LPA and EGF. These findings also suggest that the positive role of SHP2 in the MAP kinase pathway depends on its interaction with Gab1.

Successful treatment of brain tumors such as glioblastoma multiforme (GBM) is limited in large part by the cumulative dose of Radiation Therapy (RT) that can be safely given and the blood-brain barrier (BBB), which limits the delivery of systemic anticancer agents into tumor tissue. Consequently, the overall prognosis remains grim. Herein, we report our pilot studies in cell culture experiments and in an animal model of GBM in which RT is complemented by PEGylated-gold nanoparticles (GNPs). GNPs significantly increased cellular DNA damage inflicted by ionizing radiation in human GBM-derived cell lines and resulted in reduced clonogenic survival (with dose-enhancement ratio of ∼1.3). Intriguingly, combined GNP and RT also resulted in markedly increased DNA damage to brain blood vessels. Follow-up in vitro experiments confirmed that the combination of GNP and RT resulted in considerably increased DNA damage in brain-derived endothelial cells. Finally, the combination of GNP and RT increased survival of mice with orthotopic GBM tumors. Prior treatment of mice with brain tumors resulted in increased extravasation and in-tumor deposition of GNP, suggesting that RT-induced BBB disruption can be leveraged to improve the tumor-tissue targeting of GNP and thus further optimize the radiosensitization of brain tumors by GNP. These exciting results together suggest that GNP may be usefully integrated into the RT treatment of brain tumors, with potential benefits resulting from increased tumor cell radiosensitization to preferential targeting of tumor-associated vasculature.

The antineoplastic drug sorafenib ) is a multikinase inhibitor that targets the serine-threonine kinase B-Raf as well as several tyrosine kinases. Given the numerous molecular targets of sorafenib, there are several potential anticancer mechanisms of action, including induction of apoptosis, cytostasis, and antiangiogenesis. We observed that sorafenib has broad activity in viability assays in several human tumor cell lines but selectively induces apoptosis in only some lines. Sorafenib was found to decrease Mcl-1 levels in most cell lines tested, but this decrease did not correlate with apoptotic sensitivity. Sorafenib slows cell cycle progression and prevents irradiated cells from reaching and accumulating at G 2 -M. In synchronized cells, sorafenib causes a reversible G 1 delay, which is associated with decreased levels of cyclin D1, Rb, and phosphorylation of Rb. Although sorafenib does not affect intrinsic radiosensitivity using in vitro colony formation assays, it significantly reduces colony size. In HCT116 xenograft tumor growth delay experiments in mice, sorafenib alters radiation response in a schedule-dependent manner. Radiation treatment followed sequentially by sorafenib was found to be associated with the greatest tumor growth delay. This study establishes a foundation for clinical testing of sequential fractionated radiation followed by sorafenib in gastrointestinal and other malignancies. [Cancer Res 2007;67(19):9443-54]

Recent evidence indicates that the epidermal growth factor (EGF) receptor mediates a branch of lysophosphatidic acid (LPA)-induced signal transduction pathways that activate mitogen-activated protein (MAP) kinase. However, it is unclear whether the intrinsic tyrosine kinase activity of EGF receptor is involved. We previously showed that reactive oxygen species (ROS) were involved in the LPA-stimulated MAP kinase pathway. Here, we identify tyrosine phosphorylation of EGF receptor as an LPA signaling step that requires ROS. To evaluate the role of the tyrosine kinase activity of EGF receptor in the LPA-stimulated MAP kinase pathway, we examined the effects of an EGF receptor-specific tyrosine kinase inhibitor, PD158780. PD158780 potently inhibited the LPA-stimulated MAP kinase kinase 1/2 (MKK1/2) activation and EGF receptor tyrosine phosphorylation in HeLa cells, while it had no detectable effect on c-Src kinase activity. PD158780 also inhibited LPA-induced MKK1/2 activation and DNA synthesis in NIH 3T3 cells. Furthermore, we compared LPA-stimulated MKK1/2 and MAP kinase activation, transcriptional activity of the c-fos promoter, and DNA synthesis in B82L cells, which lack endogenous EGF receptor, and B82L cells expressing kinase-defective or wild-type human EGF receptor. Results obtained from analysis of these cell lines suggest that the EGF receptor tyrosine kinase contributes to the LPA-stimulated MAP kinase activation, c-fos transcription, and mitogenesis. Lysophosphatidic acid (LPA)1 is a bioactive phospholipid present in serum. LPA concentrations in serum are normally in the range of 2-20 M (1), but higher concentrations have been reported (2). LPA induces cellular responses by binding to a specific cell surface receptor(s) that is coupled to G i , G q , and G 12/13 subfamilies of heterotrimeric G-proteins (1, 3). LPA rapidly activates the mitogen-activated protein (MAP) kinase cascade, consisting of Ras, Raf-1, MAP kinase kinase (MKK or MEK) 1 and MKK 2 (MKK1/2), and MAP kinases (also called ERKs) (4 -9). It has been observed that LPA, as well as other agonists of G protein-coupled receptors, induces tyrosine phosphorylation of several signaling proteins in diverse cell types (6, 10 -15). Activation of protein tyrosine kinases, such as Src and PYK2, by agonists of G protein-coupled receptors have been reported in certain cell types (15, 16), but not others (17). The LPA-stimulated MAP kinase pathway is sensitive to certain protein tyrosine kinase inhibitors such as genistein (6). 2These observations suggest that regulation of protein tyrosine phosphorylation is an important signaling mechanism of LPA and other agonists of G protein-coupled receptors.Recently, it was reported that LPA and some other agonists of G protein-coupled receptors stimulate tyrosine phosphorylation of the epidermal growth factor (EGF) receptor (14, 15). Expression of a truncated human EGF receptor lacking the cytoplasmic domain in Rat1 cells abrogated LPA-stimulated MAP kinase activation, suggesting that the EGF receptor mediates at ...

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