WO2015154057A1 - Methods for treating cancer - Google Patents

Abstract

Various embodiments provide compositions and methods for treating glioblastoma tumors and other cancers exhibiting elevated Fn14 expression. Exemplary methods comprise applying compositions comprising aurintricarboxylic acid to the glioblastoma tumor cells to reduce migration of the cells and sensitize the cells to treatment with chemotherapeutic agents and/or radiation.

Description

Nhan Tran (Phoenix, Arizona); Harshil Dhruv (Phoenix, Arizona); and

ongwei Yin (Phoenix, Arizona)

CROSS-REFERENCES TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application

No. 61/975,632, filed April 4, 2014, and incorporates the disclosure of the application by reference in its entirety,

STATEMENT REGARDING FEDERAL RIGHTS

[0002] This invention was made with government support under CAI 77796 awarded by the National Institu tes of Health. The government has certain rights in the invention.

BACKGROUND

[0003] Glioblastoma multiforme (GBM) are cancers of the brain that originate from star-shaped astrocyte cells. GBM are the most frequently arising and most aggressive malignant brain tumors that proliferate through the central nervous system. The primary course of treatment is surgical resection of the tumor, radiation, and chemotherapy. Long-term survival of patients with GBM is compromised by the proclivity of the tumor for local tissue invasion into the surrounding normal brain tissue, which escapes surgical resection and exhibits resistance to therapies. As a result, patent with GBM general succumb to the disease within one year of diagnosis.

[0004] The tumor necrosis factor (TNF) receptor superfamily and associated ligands have a significant role in cellular processes contributing to various cancers and autoimmune diseases. The TNF receptor superfamily is involved in many developmental processes including apoptosis, regulation of immune ceil functions, and other cell-type specific responses. Fibroblast growth factor-inducible 14 (F l4), a member of the TNF receptor superfamily, is expressed at a high level in migrating glioma cells in vitro, in tissue invading glioma cells in vivo, and in GBM tissue specimens and correlates with glial tumor grade and poor clinical outcomes.

SUMMARY

[0005] Various embodiments provide compositions and methods for treating glioblastoma tumors and other cancers exhibiting elevated Fnl4 expression. Exemplary methods comprise applying compositions comprising aurmtricarboxyiic acid to the glioblastoma tumor cells to reduce migration of the cells and sensitize the cells to treatment with chemotherapeutic agents and/or radiation.

BRIEF DESCRIPTION OF THE DRAWINGS

[0006] A more complete understanding of the present technology may be derived by referring to the detailed description when considered in connection with the

9 following illustrative figures. In the following figures, like reference numbers refer to similar elements and steps throughout the figures.

Elements and steps in the figures are illustrated for simplicity and clarity and have not necessarily been rendered according to any particular sequence or scale. For example, steps that may be performed concurrently or in different order are illustrated in the figures to help to improve understanding of embodiments of the present invention.

The figures described are for illustration purposes only and are not intended to limit the scope of the present disclosure in any way. Various aspects of the present invention may be more fully understood from the detailed description and the accompanying drawing figures, wherein:

Figure 1 representatively illustrates a chemical structure of ATA;

Figure 2 representatively illustrates a schematic of T WEA -Fnl4 driven luciferase expression in a reporter cell line;

Figure 3 shows a dose response curve of inhibitory activity of ATA in HEK293 Fnl 4-NF-KB-Luc and HEK293 NF-KB-LUC cells following TWEAK and TNFa stimulation;

Figure 4 is a plot of cell viability of HEK293 FII14-NF-KB-LUC and HEK293 NF-KB-LUC cells versus concentration of ATA showing a lack of cytotoxic effects;

Figure 5 is a plot of the inhibition of migration in TWEAK induced glioma cells by ATA;

.5 [0014] Figure 6 is a plot of the inhibition of invasion in TWEAK induced glioma cells by ATA;

[0015] Figure 7 is a plot of cell viability of T98G, A172, and GBM44 cells versus concentration of ATA sho wing a lack of cytotoxic effects;

[0016] Figure 8 shows a western blot analysis of Fn-14-TWEAK signaling proteins in T98G, A172, and GBM44 glioma cells stimulated with TWEAK

(l OOng/mL) in the presence and absence of ATA (Ι Ομ,Μ);

[0017] Figure 9 shows Racl activity in T98G, A 172, and GBM44 glioma cells stimulated with TWEAK (lOOng/mL) in the presence and absence of ATA

(10μΜ);

[0018] Figure 10 shows a western blot analysis of the TRAF2 recruitment to F l4 cytoplasmic domain in ΑΓ72 and GBM44 glioma cells stimulated with TWEAK

(lOOng/mL) in the presence and absence of ATA (10μΜ);

[0019] Figure 11 shows a western blot analysis of cleaved PARP in T98G, A172, and GBM44 glioma cells treated with ATA (10μΜ), TMZ (250μΜ), or radiation;

[0020] Figure 12 shows a colon)' formation assay of T98G, A172, and GBM44 glioma cells after 24 hour treatment with ATA (10μΜ) and TMZ (250μΜ); and [0021] Figure 13 shows a colony formation assay of T98G, A172, and GBM44 glioma cells after 24 hour treatment with radiation and ATA (10μΜ).

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENT^'S

The present technology may be described in terms of functional block components and various processing steps. Such functional blocks may be realized by any number of components configured to perform the specified functions and achieve the various results. For example, methods and systems according to various aspects of the present technology may be practiced in conjunction with any number of systems and methods for diagnosing and/or treating cancer in humans and animals and the systems described are merely some exemplary applications for the technology,

[0023] The particular implementations shown and described are illustrative of the technology and its best mode and are not intended to otherwise limit the scope of the present technology in any way. For the sake of brevity, conventional manufacturing, processing, preparation, sterilization, and other functional aspects of the system may not be described in detail. Various aspects of the invention provide methods for using pharmaceutical compositions for reducing or inhibiting cancer cell migration and invasion and sensitizing the cancer cells to therapeutic treatments. A detailed description of various embodiments is provided as a specific enabling disclosure that may be generalized to any application of the disclosed systems and methods in accordance with the various described embodiments.

[0024] Gliomas, primary brain tumors that derive from glial support cells, are the most common primary tumor of the adult central nervous system. Adult gliomas of astrocytic origin (astrocytomas) comprise a spectnim of neoplasms that are generally classified by WHO standards into low-grade benign tumors (i.e. juvenile pilocytic astrocytoma, diffuse astrocytoma) and high-grade malignant tumors (i.e. anaplastic astrocytoma and glioblastoma multiforme; GBM). Patients diagnosed with grade IV GBM, the most aggressive malignant glioma, have a median survival of 9-12 months after the onset of clinical symptoms.

[0025] In general, gliomas are resistant to treatment using conventional chemotherapy and radiation therapy approaches. This is primarily due to the intrinsic propensity of glioma cells to exit the tumor core and invade the adjacent normal brain parenchyma. These migrating cells escape surgical resection and are resistant to radiation and/or cherno therapeutic agents. Migrating glioma cells also exhibit decreased susceptibility to pro-apoptotic agents. The migrating glioma cells may travel over long distances, frequently along blood vessel and fiber tracts, and then initiate secondary tumor growth at their final destination. This distinguishing invasive ability is not shared by nongliai cells that metastasize from other primar tumor sites (e.g. breast) to brain tissue.

[0026] The elucidation of the mechanism for migration activation of glial neoplasms may comprise a variety of cellular signals that promote a cascade of cellular responses such as cell migration, ititegrin-mediated cell attachment to extracellular matrix (ECM) molecules, cell-cell adhesion, the production and secretion of ECM-degrading enzymes, and/or cellular motility. See Pilkington et al Int. J. Dev. Neurosci. 17:613-623 (1999), Pilkington et al. Curr. Neurol. Neurosci. Rep. 1 : 225-232 (2001), Giese et al. Clin Exp Metastasis 12: 405-415 (1994), Gnmiet et al. Cancer Res. 56: 1939-1947 (1996), Berens et al. Cancer Res. 54: 3897-3904 (1994).

[0027] Various embodiments of the present technology provide compositions and methods for treating glioblastoma tumors and other cancers in a subject. The subject includes any human patient or non-human mammal, including for example: a primate, cow, horse, pig, sheep, goat, dog, cat, or rodent, capable of developing cancer including human patients that are suspected of having cancer, that have been diagnosed with cancer, or that have a family histor of cancer. Methods of identifying subjects suspected of having cancer include but are not limited to: physical examination, family medical history, subject medical history including exposure to environmental factors, biopsy, or any of a number of imaging technologies such as ultrasonography, computed tomography, magnetic resonance imaging, magnetic resonance spectroscopy, or positron emission tomography.

[0028] The cancer ceils include any cells derived from a tumor, neoplasm, cancer, precancer, cell line, malignancy, or any other source of cells that have the potential to expand and grow to an unlimited degree. Cancer cells may be derived from naturally occurring sources or may be artificially created. Cancer cells may also be capable of invasion into other tissues and metastasis. Cancer cells further encompass any malignant cells that have invaded other tissues and/or metastasized. One or more cancer cells in the context of an organism may also be called a cancer, tumor, neoplasm, growth, malignancy, or any other term used in the art to describe ceils in a cancerous state.

[0029] Examples of cancers that may be treatable with compositions and methods of the present invention may comprise solid tumors such as fibrosarcoma, myxosarcoma, iiposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lyrnphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon cancer, colorectal cancer, kidney cancer, pancreatic cancer, bone cancer, breast cancer, ovarian cancer, prostate cancer, esophageal cancer, stomach cancer, oral cancer, nasal cancer, throat cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcmoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, cervical cancer, uterine cancer, testicular cancer, small cell lung carcinoma, bladder carcinoma, lung cancer, epithelial carcinoma, glioma, glioblastoma multiforme, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma, skin cancer, melanoma, neuroblastoma, and retinoblastoma.

Additional cancers may include blood borne cancers such as acute lymphoblastic leukemia ("ALL,,"), acute lymphoblastic B-cell leukemia, acute lymphoblastic T-cell leukemia, acute myeloblastic leukemia ("AML"), acute promyelocyte leukemia ("APL"), acute monobiastic leukemia, acute erythroleukemic leukemia, acute megakaryoblastic leukemia, acute myelomonocytic leukemia, acute nonlyrnphocyctic leukemia, acute undifferentiated leukemia, chronic myelocytic leukemia ("CML"), chronic lymphocytic leukemia f'Ol .[.'^" )„ hairy ceil leukemia, multiple myeloma, lymphoblastic leukemia, myelogenous leukemia, lymphocytic leukemia, myelocytic leukemia, Hodgkin's disease, non- Hodgkin's Lymphoma, Waldenstrom's macroglobulinemia, Heavy chain disease, and Polycythemia vera.

Subjects with glioblastoma or other brain tumors may exhibit particular symptoms that signify the presence of a tumor or other growth in the brain. Such symptoms may include headache, seizures, mental or personality changes, mass effect, or one of a number of focal or localized systems including ringing or buzzing sounds, hearing loss, loss of coordination, reduced sensation, weakness or paralysis, difficulty with walking or speech, difficulty keeping balance, decreased muscle control, or double vision. The subject may display one or more different brain tumor types including acoustic neurinoma, astrocytoma, ependyoma, glioblastoma multiforme, meningioma, metastatic tumors originating from another tumor type, mixed glioblastoma, oligodendroglioblastoma, or pineal region tumor.

Glioblastoma tumors and/or other cancers that may be susceptible to treatment with the present compositions and methods. Treatment of the cancer is the practice of any method, process, or procedure with the intent of halting, inhibiting, slowing or reversing the progression of a disease, disorder or condition, substantially ameliorating clinical symptoms of a disease disorder or condition, or substantially preventing the appearance of clinical symptoms of a disease, disorder or condition, up to and including returning the diseased entity to its condition prior to the development of the disease.

The susceptible glioblastoma tumors and/or other cancers may exhibit elevated gene expression of the fibroblast growth factor-inducible 14 (Fill 4) gene. Fnl4 is a member of the tumor necrosis factor (TNF) superfamily of receptors and is characterized as a type-i transmembrane receptor lacking a cytoplasmic death domain. See Winkles et ai. J. Biol. Chem. 274: 33166-33176 (1999), Winkles et al. Am. J. Pathol. 156: 1253-1261 (2000), and Fanslow et. al . Immunity 15: 837- 846 (2001 ). Fn!4 has been shown to be up-regulated in migration-stimulated glioma cells in vitro and to have minimal to absent expression in normal brain tissue. See Tran et al. Am. J. Pathol. 162: 1316 (2003). Additionally, Fnl4 expression has been demonstrated to increase with histopathological grade of glial neoplasms. See Id. at 1317.

[0034] Activation of Fnl4 signaling by the Fnl4 ligand TNF-like weak inducer of apoptosis (TWEAK) has been demonstrated to stimulate migration of glioma cells and causes up-regulation of expression of Fnl4. See Berens et al. Cancer Res. 66(1): 9536- 37 (2006). TWEAK is a type I membrane protein and member of the tumor necrosis factor (TNF) ligand superfamily. TWEAK has been demonstrated to stimulate glioma cell invasion in vivo via induction of Fill expression levels via the Racl IKKp NF-KB pathway. Id. at 9538-39. See also U.S. Patent Publication No. 2015/0017263, the disclosure of which is incorporated herein by reference.

[0035] Various embodiments of the present technology demonstrate that aurintricarbox lic acid (ATA) is a specific inhibitor of TWEAK-Fnl 4-NF- B signaling. Referring to the structure of ATA illustrated in Figure 1 , ATA is a cell permeable polyanionic, poiyaromatic compound that may be used to inhibit cellular processes, such as apoptosis, that may involve the formation of protein- nucleic acid complexes. As discussed below, ATA suppresses TWEAK induced chemotactic migration of human glioma cell lines and displays no general cytotoxicity. ATA also sensitizes glioma cells to cell death from treatment with radiation and/or chemotherapeutic agents.

ATA suppression of TWEAK-Fn l4 mediated NF-κΒ activation

[0036] Referring to Figure 2, a schematic of TWEAK mediated Fn l4 cell signaling is illustrated in the context of a firefly lueiferase expression system. In this expression system, firefly lueiferase is a bioluminescent protein whose expression is driven by the NF-κΒ transcription factor. Binding of TWEAK to Fnl 4 promotes trimerization of Fnl4 in the cell membrane and association of TNF Receptor (TNFR)-associated factor (TRAP). Subsequent signal pathway processes activates NF- Β, which translocates to the nucleus to drive NF-κΒ dependent gene expression of firefly lueiferase. Bioluminescence of firefly lueiferase is quantified using standard protocols. See Promega Lueiferase Assay System.

[0037] Figure 3 depicts a dose response curve showing the inhibitory activity of

ATA in HEK293 FTI14-NF- B-LUC and HEK293 NF-KB-LUC ceils following TWEAK and TNFa stimulation, respectively. As the concentration of ATA increases, the relative light units (RLIJ) emitted by the firefly lueiferase decreases, indicating the specific inhibition of TWEAK-Fnl4 driven NF- Β activation with ICso concentration of 0.6 μΜ of ATA.. AT A did not show any inhibition of NF-KB activity downstream of T Fa stimulation. Additional ly, ATA did not demonstrate any cytotoxic effects to the HEK293 Fnl 4-NF-KB-Luc and HEK293 NF-KB-LUC cells, as shown in Figure 4. A CellTiterGlo® assay measured the percent cell viability with increasing AT A concentration. The absence of general cytotoxicity indicates that the inhibitory effect of ATA on TWEAK-Fnl4 signaling is due to a specific pharmacological effect by ATA.

ATA s 3 pression of glioma cell migration and in vasion

[0038] In some aspects of the present technology, ATA suppresses TWEAK induced chemotactic migration of glioma cells (T98G and A 172) and displays no general cytotoxicity. Normal, noncancerous cells do not migrate and any inhibition of glioma cell migration is significant. Studies on glioma cells have demonstrated a strong correlation between migration rate in vitro and invasive behavior in vivo. See Sondak et al. Nature 406:536-540 (2000) and Silbergeld et al Cancer 75: 2904-2909 (1995).

[0039] Figure 5 is a plot showing an assay measuring cellular migration of A172,

GBM44, and T98G glioma cells. Figure 6 is a plot showing an assay measuring cellular invasion of these glioma cells. Cell migration activity includes any mode through which a ceil may move in two-dimensional or three-dimensional space. Such migration includes movement through the use of pseudopodia including the adhesion of pseudopodia to a surface, a flagellum, a cilium, acts of amoeboid movement, extravasation, myosin-actin interactions, microtubule extension, or any other process through which a cell moves itself from one place to another or changes its morphology. In one aspect of the invention, cell migration activity is measured through cell adhesion. Using adhesion, cell migration activity may be measured by cell-cell aggregation, monolayer radial migration, including adhesion to a cell matrix comprising laminin, BSA or any other cell matrix component, three dimensional spheroid dispersion, or any other method that measures adhesion based cellular migration in space. Migration activity may be measured by any method that detects that a cell has moved from one place to another or has changed its morphology. Suc methods include flow cytometry, capillary electrophoresis, visual examination by light, fluorescence, or electron microscopy, or any such method known in the art or yet to be developed. Inhibitors of ceil migration activity are agents that disrupt any molecular or cellular process involved in cell mi gration activity .

In the assay of Figure 5, the ceils were added to the top well of a modified transweli chamber pre-coated with collagen in serum-free DMEM + 0.1% BSA to assess migration, or, as shown in Figure 6, pre-coated with lOmg/mL growth factor-free Matrigel to assess invasion. TWEAK alone (100 ng/ml.) or TWEAK, and ATA combination (10 uM) was added to the lower wells and the number of cells migrated observed in the bottom chamber was quantitated after 5 hrs. Values are mean ± standard deviation of triplicate measurements (*, p < 0.01).

The first column of Figure 5 and 6 under eac cell line shows the baseline amount of migration or invasion of each cell line in the absence of TWEAK and ATA. The second column under each cell line shows the elevated amount of migration and invasion of each ceil line as stimulated by the addition of TWEAK. The third and fourth columns under each cell line show the reduction of migration and invasion of each cell line upon treatment with ATA. Accordingly, ATA suppresses T WEA induced migration and invasion of glioma cells.

Referring to Figure 7, the absence of a cytotoxic effect of ATA on glioma cells was ruled out as contributing to the decrease of migration and in vasion of the glioma cells in Figures 5 and 6 via a cell viability assay. Ceil viability was assessed by quantifying metabolic activity of the glioma cells after ATA treatment . The captioned glioma cells were seeded in 96-vvell plates and incubated for 24 hours. A vehicle control (DMSO) or ATA at the indicated concentrations were added to each well. After 72 hours of incubation, the viability of the cells was measured using CeilTiterGio® assay kit. Values are mean ± standard deviation of six separate measurements. As the plot indicates, the viability of each cell line was not affected by increasing concentrations of ATA, indicating that ATA- induced reduction of migration and invasion of the glioma cells in Figures 5 and 6 were not due to a reduction of cell viability.

ATA mediated suppression of TWEAK-Fnl4 cell signaling in glioma cells [0043] Referring to Figure 8, ATA is shown to mediate suppression of TWEAK-

Fnl4 cell signaling in glioma cells. T98G, A172, and GBM44 glioma cells were stimulated with TWEAK (100 iig/mL) in presence or absence of ATA (10 μΜ) for 10 min and lysed. Western blot analysis was performed using Phospho NF-KB (S536), Total NF-κΒ, Pan Phospho Src (Y416), Total Src, Phospho AKT (Y473), and Total AKT (a-Tubulin used as loading control).

[0044] The first column of Figure 8 under each cell line shows a baseline minimal phosphorylation of NF- Β, AKT, and Src in the absence of TWEAK and ATA. The second column under each cell line shows the phosphorylation of each of NF- KB, AKT, and Src in the presence of TWEAK stimulation. The third column under each cell line shows the baseline minimal phosphorylation of NF-κΒ, AKT, and Src in the presence of ATA.. However, as shown in the fourth column under each ceil line, ATA prevents the phosphorylation of each of NF-κΒ, AO', and Src in the presence of TWEAK, Accordingly, ATA suppresses Fn 14 -TWEAK induced survival and migration signaling.

Referring to Figure 9, the inhibition of migration and invasion of glioma cells by ATA is further supported by the ability of ATA to suppress TWEAK induced Racl activation in glioma ceils, Racl is a member of the Rho family of smal l GTPases that are important mediators of the reorganization of cellular actin cytoskeleton that is integral to cell migration. See Burridge et al. Science 302: 1704-9 (2003), Marshall et al. Nat. Rev. Cancer 2: 133-42 (2002), and Hall et al. J. Cell Biol 144: 1235-44 (1999). Specifically, Racl stimulates actin assembly and the formation of lameilipodia. See Hall et al. Dev. Biol. 265: 23-32 (2004).

In Figure 9, T98G and A 172 glioma cells were stimulated with TWEAK (100 ng/^'mL) in presence or absence of ATA (10 μΜ) for 10 minutes and lysed. Racl activity was determined using a Racl activity assay kit as previously described. See Tran et al., Mol Cancer Res 8: 1560 (2010). The first column under each cell line shows a baseline level of active Rac I in the absence of ATA and TWEAK. The second column shows that TWEAK stimulates the Racl activation. The third column shows that ATA does not stimulate Racl . The fourth column shows that ATA. inhibits the ability TWEAK to activate Racl .

Figure 10 is a western blot for Fnl4 and TRAF2 (as a loading control) showing that ATA suppresses TWEAK induced TRAF2 recruitment to the Fnl4 cytoplasmic domain in glioma cells, A172 and GBM44 glioma cells were stimulated with TWE AK (100 ng/mL) in presence or absence of AT A (10 μΜ) for 2 min and lysed. Cell lysates were immunopreeipitated using TRAF2 antibody or IgG control antibody and western blotted for F l4 and TRAF2 (as a loading control). The lane labeled WB:Fnl4 shows that a TRAF2 immunoprecipitate is bound to Fn!4 in the presence of TWEAK (second lane under each cell line). As illustrated in Figure 2, above, TWEAK induces trimerization of Fnl4, which recruits TRAF2 to the cytoplasmic domain of Fnl4. However, in glioma ceils stimulated with TWEAK, a TRAF2 immunoprecipitate is not bound to F l 4 in the presence of ATA (fourth lane under each cell line). This observation indicates that ATA suppresses TRAF2 recmitment to Fnl4 cytoplasmic domain downstream of TWEAK activation.

[0048] In summary, ATA specifically suppresses TWEAK~Fnl4 induced migration and invasion of glioma cells. Additionally, ATA suppresses TWEAK- Fnl4 induced Racl activation, which plays important role in migration and invasion of cells. Further, ATA acts upstream of TRAF2 for inhibiting TWEAK- Fnl4 signaling axis.

Sensitization of glioma cells to chemotherapeutic agents and radiation therapy

[0049] Glioblastoma tumors have a propensity to become resistant to chemotherapeutic agents and radiation therapy. Glioblastoma tumors may develop resistance to apoptosis, which promotes tumor development and progression. Temozolomide is a chemotherapeutic agent that causes apoptosis and is rendered ineffective against glioblastoma tumors due to the apoptosis resistance of these cells. See Spruce et ai, Cancer Res 64, 4875-4886 (2004) and Decaestecker et al, Med Res Rev 27, 149-176 (2007). [0050] Referring to Figure 11, poly (ADP-ribose) polymerase (PARP) is cleaved during apoptosis by caspase-3. See Sraulson et al., J. Biol, Chem. 274: 22932 (1999). A western blot using ant.i-PA.RP cleavage fragment antibodies is an indicator of apoptosis. T98G, A172, and GBM44 glioma cells were treated with: ATA (10 μΜ), TMZ (250 μΜ), ATA + TMZ, Radiation (2 Gy), or ATA + Radiation in presence or absence of TWEAK (100 ng/mL) for 48 hours and iysed. Western blot analysis was performed using anti-cleaved PARP antibodies and a- Tubulin as loading control).

[0051] The western blot analysis shows that TWEAK confers chemo- and radiation resistance to glioma cells. Specifically, in the presence of TWEAK, the amount of cleave PARP is reduced after treatment with TMZ and radiation suggesting a protective effect by TWEAK (Lanes 6 and 14, respectively). However, in presence of ATA, TWEAK-Fnl4 signaling is inhibited and glioma cells show increased cleaved PARP in the presence of TWEAK after TMZ and radiation therapy, as compared to absence of ATA (Lane 8 and 16, respectively).

[0052] Referring to Figures 12 and 13, the efficacy of ATA in sensitizing glioma cells to chemotherapy and radiotherapy, respectively, was assessed by colony formation assay. A172, GBM44, and T98G glioma cells were treated with ATA (10 μΜ), TMZ (250 μΜ), ATA + TMZ, Radiation (2Gy), or Radiation + ATA in presence or absence of TWEAK (100 ng mL) for 24 hours. After 24 hours of dmg treatment, cells were trypsinized and 250 ceils were seeded in triplicates in 35 mm dishes and allowed to form colonies. At the end of the assay, cells were fixed in paraformaldehyde and stained with crystal violet. The number of colonies were counted and presented here as bar graph. Values are mean ± standard deviation of three separate measurements.

Colony formation analysis demonstrates that TWEAK confers chemo- and radiation resistance to glioma cells, as in the presence of TWEAK glioma cells do not show any difference in clonogenicity when compared to non-treated cells (Lanes 1 and 4 under each ceil line). However, in the presence of ATA, TWEAK- Fnl4 signaling is inhibited and glioma cells show significantly lower clonogenicity, as compared to non-treated cells (Lanes 8 and 1 under each cell line). Accordingly, ATA treatment specifically suppresses TWEAK-Fnl4 signaling and subsequently sensitizes glioma cells to TWEAK induced chemo- and radiation protection.

Pharmaceutical Formulations

^'The present compositions encompass pharmaceutical compositions comprising one or more active phannaceuticai ingredients, such as, but riot necessarily limited to ATA, These pharmaceutical compositions may comprise suitable carriers, such as pharmaceutical iy acceptable excipients including buffers, which are well known in the art. The concept of a pharmaceutical composition including an active pharmaceutical agent, such as ATA, also encompasses the active pharmaceutical agent without any other additive.

Such phannaceuticai compositions may take any physical form necessary depending on a number of factors including the desired method of administration and the physicochemicai and stereochemical form taken by the active pharmaceutical agent or pharmaceutically acceptable salts thereof. Such physical forms include a solid, liquid, gas, sol, gel, aerosol, or any other physical form now known or yet to be disclosed. The physical form of the pharmaceutical compositions may affect the route of administration and one skilled in the art. would know to choose a route of administration that takes into consideration both the physical form of the active pharmaceutical agent and the disorder to be treated.

[0056] Pharmaceutical compositions that include the active pharmaceutical agent may be prepared using methodology well known in the pharmaceutical art. A pharmaceutical composition that includes the active pharmaceutical agent may include a second effective compound of a distinct chemical formula from the active pharmaceutical agent. For example, in some embodiments, the pharmaceutical composition can include a combination of any of the aforementioned or later disclosed active pharmaceutical agents described in this instant application. This second effective compound may have the same or a similar molecular target as the target or it may act upstream or downstream of the molecular target of the active pharmaceutical agent wit regard to one or more biochemical pathways.

[0057] Pharmaceutical compositions including the active pharmaceutical agent may include materials capable of modifying the physical form of a dosage unit. The dosage unit is an individual or single use dose of the pharmaceutical composition. In one nonlimiting example, the pharmaceutical composition includes a material that forms a coating that holds in the pharmaceutical composition. Materials that may be used in such a coating, include, for example, sugar, shellac, gelatin, or any other inert coating agent. Pharmaceutical compositions including the active pharmaceutical agent may be prepared as a gas or aerosol Aerosols encompass a variety of systems including colloids and pressurized packages. Delivery of a pharmaceutical composition in this form may include propulsion of the pharmaceutical composition including the active pharmaceutical agent through use of liquefied gas or other compressed gas or by a suitable pump system. Aerosols may be delivered in single phase, bi-phasic, or tri- phasic systems.

In some aspects of the technology, the pharmaceutical composition may comprise an active pharmaceutical agent in the form of a solvate. Such solvates may be produced by the dissolution of the active pharmaceutical agent in a pharmaceutically acceptable solvent. Pharmaceutically acceptable solvents include any mixtures of more than one solvent. Such solvents may include pyridine, chloroform, propan-1-οί, ethyl oieate, ethyl lactate, ethylene oxide, water, ethanol, and any other solvent that delivers a sufficient quantity of the disclosed compound to treat the affliction without serious complications arising from the use of the solvent in a majority of patients.

Pharmaceutical compositions may also include a pharmaceutically acceptable carrier. Carriers include any substance that may be administered with the disclosed pharmaceutical composition with the intended purpose of facilitating, assisting, or helping the administration or other delivery of the pharmaceutical composition. Carriers may include any liquid, solid, semisolid, gel, aerosol or anything else that may be combined with the pharmaceutical composition to aid in its administration. Examples include diluents, adjuvants, excipients, water, oils (including petroleum, animal, vegetable or synthetic oils.) Such carriers include particulates such as a tablet or powder, liquids such as an oral syrup or injectable liquid, and inhalable aerosols. Further examples include saline, gum acacia, gelatin, starch paste, talc, keratin, colloidal silica, and urea. Such carriers may further include binders such as ethyl cellulose, carboxymethylcellulose, microcrystalline cellulose, or gelatin; excipients such as starch, lactose or dextrins; disintegrating agents such as alginic acid, sodium alginate, Primogel, and com starch; lubricants such as magnesium stearate or Sterotex; glidants such as colloidal silicon dioxide; sweetening agents such as sucrose or saccharin, a flavoring agent such as peppermint, methyl salicylate or orange flavoring, or coloring agents. Further examples of earners include polyethylene glycol, cyclodextrin, oils, or any other similar liquid carrier that may be formulated into a capsule. Still further examples of carriers include sterile diluents such as water for injection, saline solution, physiological saline. Ringer's solution, isotonic sodium chloride, fixed oils such as synthetic mono or digylcerides, polyethylene glycols, glycerin, cyclodextrin, propylene glycol or other solvents; antibacterial agents such as benzyl alcohol or methyl paraben; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethyienediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose, thickening agents, lubricating agents, and coloring agents.

The pharmaceutical composition including the active pharmaceutical agent may take any of a number of formulations depending on the pbysicochemical form i of the composition and the type of administration. Such forms include solutions, suspensions, emulsions, tablets, pills, pellets, capsules, capsules including liquids, powders, sustained-release formulations, directed release formulations, lyophylates, suppositories, emulsions, aerosols, sprays, granules, powders, syrups, elixirs, or any other formulation now known or yet to be disclosed. Additional examples of suitable pharmaceutical carriers are described in "Remington's Pharmaceutical Sciences" by E. W. Martin, hereby incorporated by reference in its entirety.

Administration of pharmaceutical compositions

[0062] Various embodiments of the present pharmaceutical compositions may be administered in a therapeutically effective amount to the subject. For example, in one embodiment, the therapeutically effective amount may be up to about 50 milligrams of ATA per kilogram (50 mg/kg) of the subject's body weight. Determination of an effective amount of the pharmaceutical compositions is within the capability of those skilled in the art, especially in light of the detailed disclosure provided herein.

[0063] The effective amount of a pharmaceutical composition used to affect a particular purpose as well as the pharmacologically acceptable dose may be detennined by toxicity, excretion, and overall tolerance may be determined in cell cultures or experimental animals by pharmaceutical and toxicological procedures either known now by those skilled in the art or by any similar method yet to be disclosed. One example is the determination of the ICso (half maximal inhibitory concentration) of the pharmaceutical composition in vitro in cell lines or target zz molecules. Another example is the determination of the LD50 (lethal dose causing death in 50% of the tested animals) of the pharmaceutical composition in experimental animals.

[0064] The exact techniques used in determining a therapeutically effective amount may depend on factors such as the type and physical/chemical properties of the pharmaceutical composition, the property being tested, and whether the test is to be performed in vitro or in vivo. The determination of an effective amount of a pharmaceutical composition will be well known to one of skill in the art who will use data obtained from any tests in making that determination. Determination of an effective amount of disclosed compound for addition to a cancer cell also includes the determination of an effective therapeutic amount, including the formulation of an effective dose range for use in vivo, including in humans.

[0065] The therapeutically effective dose of the pharmaceutical composition may be that amount effective to prevent occurrence of the symptoms of a disorder or to treat some symptoms of the disorder from which the patient suffers. The therapeutically effective dose may also include an effective amount, a therapeutic amount, or any amount sufficient to elicit the desired pharmacological or therapeutic effects, thus resulting in effective prevention or treatment of the disorder. Thus, when treating a patient with glioblastoma, the therapeutically effective amount of the pharmaceutical composition is an amount sufficient to slow, or arrest the progression, migration, metastasis, growth, or development of the tumor with the result that life is extended. Prevention includes a delay in onset of symptoms. Treatment includes a decrease in the symptoms associated with the disorder or an amelioration of the recurrence of the symptoms of the disorder. A pharmacologically acceptable dose encompasses any dose that may be administered to a patient that will not be lethal to the patient or cause effects that threaten the health or the life of the patient,

A therapeutically effective amount further includes the prevention of progression of the cancer to a neoplastic, malignant or metastatic state. Such preventative use is indicated in conditions known or suspected of preceding progression to cancer, in particular, where non- or precancerous cell growt consisting of hyperplasia, metaplasia, or most particularly, dysplasia has occurred (for review of such abnormal growth conditions, see Robbins and Angell, 1976, Basic Pathology, 2d Ed., W. B. Saunders Co., Philadelphia, pp. 68-90, incorporated by reference). Hyperplasia is a form of controlled cell proliferation involving an increase in cell number in a tissue or organ, without significant alteration in structure or activity. For example, endometrial hyperplasia often precedes endometrial cancer and precancerous colon polyps often transform into cancerous lesions. Metaplasia is a form of controlled cell growth in which one type of adult or fully differentiated cell substitutes for another type of adult cell. Metaplasia can occur in epithelial or connective tissue cells. A typical metaplasia involves a somewhat disorderly metaplastic epithelium. Dysplasia is frequently a forerunner of cancer, and is found mainly in the epithelia; it is the most disorderly form of non-neoplastic cell growth, involving a loss in individual cell uniformity and in the architectural orientation of cells, Dysplastic cells often have abnormally large, deeply stained nuclei, and exhibit pleornorphism. Dysplasia characteristically occurs where there exists chronic irritation or inflammation, and is often found in the cervix, respiratory passages, oral cavity, and gall bladder.

[0067] Alternatively or in addition to the presence of abnormal cell growth characterized as hyperplasia, metaplasia, or dysplasia, the presence of one or more characteristics of a transformed phenotype or of a malignant phenotype, displayed in vivo or displayed in vitro by a cell sample derived from a patient can indicate the desirability of prophylactic/therapeutic administration of the pharmaceutical composition that includes the compound. Such characteristics of a transformed phenotype include morphology changes, looser substratum attachment, loss of contact inhibition, loss of anchorage dependence, protease release, increased sugar transport, decreased serum requirement, expression of fetal antigens, disappearance of the 250,000 dalton cell surface protein, etc. Further examples include leukoplakia, featuring a benign-appearing hyperplastic or dyspiastic lesion of the epithelium, or Bow en's disease, a carcinoma in situ. Both of these are precancerous lesions indicative of the desirability of prophylactic intervention. In another example, fibrocystic disease including cystic hyperplasia, mammary dysplasia, adenosis, or benign epithelial hyperplasia is indicates desirability of prophylactic intervention.

[0068] Various embodiments of the present technology may be used in, for example, in vitro, ex vivo, and/or in vivo therapeutic methods. In various embodiments of the present technology, the pharmaceutical composition comprising ATA may be administered to the subject with cancer in conjunction with any suitable treatment modality. Such treatment modalities include but are not limited to, radiotherapy, chemotherapy (e.g., conventional chemotherapy, including the use of compounds such as temozolomide), surgery, immunotherapy, cancer vaccines, radioimmunotherapy, treatment wit a pharmaceutical composition other than those which include ATA, or any other method that effectively treats cancer in combination with the pharmaceutical composition now known or yet to be disclosed. Combination therapies may act synergisticaily. That is, the combination of the two therapies is more effective than either therapy administered alone. This results in a situation in which lower dosages of bot treatment modality may be used effectively. This in turn reduces the toxicity and side effects, if any, associated with the administration either modality without a reduction in efficacy.

In another aspect of the invention, the pharmaceutical composition may be administered in combination with a therapeutically effective amount of radiotherapy. The radiotherapy may be administered concurrently with, prior to, or following the administration of the pharmaceutical composition comprising ATA. The radiotherapy may act additively or synergisticaily with the pharmaceutical composition. This particular aspect of the invention may be most effective in cancers known to be responsive to radiotherapy. Cancers known to be responsive to radiotherapy include, but are not limited to, Non-Hodgkm's lymphoma, Hodgkin's disease, Ewing's sarcoma, testicular cancer, prostate cancer, ovarian cancer, bladder cancer, laiynx cancer, cervical cancer, nasopharynx cancer, breast cancer, colon cancer, pancreatic cancer, head and neck cancer, esophageal cancer, rectal cancer, small-cell lung cancer, non-small ceil lung cancer, brain tumors, other CNS neoplasms, or any other such tumor now known or yet to be disclosed.

[0070] The pharmaceutical compositions of the present technology may be used in conjunction with any suitable chemotherapeutic agents/compositions such as cis- diamminedichioro platinum (IS) (cisplatin), doxorubicin, 5-fluorouracil, taxol, and topoisomerase inhibitors such as etoposide, teniposide, irinotecan and topotecan. For example, the chemotherapeutic agents might also include TROY inhibitors, Pyk2 inhibitors, Racl inhibitors, Dock! 80 inhibitors, Dock? inhibitors, temozolomide and bevacizumab. Still other pharmaceutical compositions include antiemetic compositions such as metociopromide, domperidone, prochlorperazine, promethazine, chlorpromazme, trimethobenzamide, ondansetron, granisetron, hydroxyzine, acethy!leucine monoethanolamine, alizapride, azasetron, benzquinamide, bietanautine, bromopride, buclizine, clebopride, cyclizine, dinienliydriiiate, diplienidol, dolasetroii, meclizine, methallatal, metopimaziiie, nabilone, oxypemdyl, pipamazine, scopolamine, sulpiride, tetrahydrocannabinols, thiethylperazine, thioproperazine and tropisetron.

[0071] In some embodiments, the pharmaceutical compositions that may be administered to the subject in combination with hematopoietic colony stimulating factors. Examples of hematopoietic colony stimulating factors include, but are not limited to, filgrastim, sargramostim, molgramostim and epoietin alia. Alternatively, the pharmaceutical composition may be used in combination with an anxiolytic agent. Examples of anxiolytic agents include, but are not limited to, buspirone, and benzodiazepines such as diazepam, lorazepam, oxazapam, chiorazepate, clonazepam, chlordiazepoxide and alprazolam,

[0072] The pharmaceutical compositions may further be used in combination with analgesic agents. Such agents may be opioid or non- opioid analgesic. Non- limiting examples of opioid analgesics inlcude morphine, heroin, hydromorphone, hydroeodoiie, oxyniorphone, oxycodone, metopon, apomorphiiie, normorphine, iorpiiiu . buprenorphine, meperidine, lopermide, anileridme, ethoheptazine, piminidine, betaprodine, diphenoxylate, fentanil, sufentanil, alfentanil, remifentanil, levorphanol, dextromethorphan, phenazocine, pentazocine, cyclazocine, methadone, isomethadone and propoxyphene. Suitable non-opioid analgesic agents include, but are not limited to, aspirin, celecoxib, rofecoxib, die!ofinac, diflusinal, etodolac, fenoprofen, flurbiprofen, ibuprofen, ketoprofen, indomethacin, ketorolac, meclofenamate, mefanamic acid, nabumetone, naproxen, piroxicam, sulindac or any other analgesic now known or yet to be disclosed.

[0073] Pharmaceutical compositions of the present technology encompass inhibitors of cell migration activity and inhibitors of effector recruitment activity. Inhibition encompasses any action that hinders, from any detectable level up to and including complete inactivation, the progression of a biological process. Such biological processes include expression of a gene or activities of a gene product, progression of a disease, normal and abnormal metabolic activities, interactions between entities within an organism, or interactions between one organism and another. Further nonlimiting examples of biological processes include development, death, maturation, infection, pain, apoptosis, or homeostasis, Inhibition includes actions that silence or repress the expression of a gene. Inhibition also includes actions that hinder the activity of the RNA product, protein product, or postranslationally modified protein product of a gene. Inhibition may be effectuated through a single agent that inactivates a single gene or gene product, by a single agent that mactivates a combination of more than one gene or gene product, a combination of agents that inactivates a single gene or gene product or a combination of agents that mactivates a combination of more than one gene or gene product.

[0074] Inhibition may be effectuated directly by an agent that directly causes the inhibition of a biological process or by agents that trigger one or more different biological processes to effectuate the inhibition of the first biological process. Agents that cause inhibition may also be called inhibitors. Examples of inhibitors mclude compositions such as compounds that trigger RNAi silencing such as microRNA or siRNA, small molecular compounds, proteins such as soluble receptors or antibodies or any fragment thereof, including an Fab, F(ab)2, Fv, scFv, Fc, phage displ ay antibody, peptibody or any other composition of matter that may inactivate or hinder a biological process. Further nonlimiting examples of inhibitors include X-rays, UV rays, visible light including laser light, and sound.

[0075] Methods of administration of the pharmaceutical compositions and treatment modalities include, but are not limited to, oral administration and parenteral administration. Parenteral administration includes, but is not limited to intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, sublingual, intranasal, intracerebral, iratra ventricular, intrathecal, intravaginal, transdermal, rectal, by inhalation, or topically to the ears, nose, eyes, or skin. Other methods of administration include but are not limited to infusion techniques including infusion or bolus injection, by absorption through epithelial or mucocutaneous linings such as oral mucosa, rectal and intestinal mucosa. Compositions for parenteral administration may be enclosed in ampoule, a disposable syringe or a multiple-dose vial made of glass, plastic or other material.

[0076] Administration may be systemic or local. Local administration is administration of the disclosed compound to the area in need of treatment. Examples include local infusion during surgery; topical application, by local injection; by a catheter; by a suppository; or by an implant. Administration may be by direct injection at the site (or former site) of a cancer, tumor, or precancerous tissue or into the central nervous system by any suitable route, including intraventricular and intrathecal injection. Intraventricular injection can be facilitated by an intraventricular catheter, for example, attached to a reservoir, such as an Ommaya reservoir. Pulmonary administration may be achieved by any of a number of methods known in the art. Examples include use of an inhaler or nebulizer, formulation with an aerosolizing agent, or via perfusion in a fluorocarbon or synthetic pulmonary surfactant. The disclosed compound may be delivered in the context of a vesicle such as a liposome or any other natural or synthetic vesicle,

[0077] The pharmaceutical composition may be formulated so as to be administered by injection by dissolving the active pharmaceutical agent with water so as to form a solution. In addition, a surfactant may be added to facilitate the formation of a homogeneous solution or suspension. Surfactants include any complex capable of non-eovalent interaction with the active pharmaceutical agent so as to facilitate dissolution or homogeneous suspension of the active pharmaceutical agent.

[0078] Pharmaceutical compositions may be prepared in a form that facilitates topical or transdermal administration. Such preparations may be in the form of a liquid solution, cream, paste, lotion, shake lotion, powder, emulsion, ointment, gel base, transdermal patch or iontophoresis device. Examples of bases used in such compositions include opetrolatum, lanolin, polyethylene glycols, beeswax, mineral oil, diluents such as water and alcohol, and emulsifiers and stabilizers, thickening agents, or any other suitable base now known or yet to be disclosed.

[0079] The technology further encompasses kits that facilitate the administration of the pharmaceutical composition to the diseased subject. An example of such a kit includes one or more unit dosages of the pharmaceutical composition. The unit dosage would be enclosed in a preferably sterile container and would be comprised of the disclosed compound and a pharmaceutically acceptable carrier. In another aspect, the unit dosage would comprise one or more lyophilates of the pharmaceutical composition. In this aspect of the invention, the kit may include another preferably sterile container enclosing a solution capable of dissolving the lyophilate. However, such a solution need not be included in the kit and may be obtained separately from the lyophilate. In another aspect, the kit may include one or more devices used in administrating the unit dosage. Examples of such devices include, but are not limited to, a syringe, a drip bag, a patch or an enema. In some aspects of the invention, the device comprises the container that encloses the unit dosage.

[0080] In the foregoing description, the technology has been described with reference to specific exemplar ' embodiments. Various modifications and changes may be made, however, without departing from the scope of the present technology as set forth. The description and figures are to be regarded in an illustrative manner, rather than a restrictive one and all such modifications are intended to be included within the scope of the present technology. Accordingly, the scope of the technology should be determined by the generic embodiments described and their legal equivalents rather than by merely the specific examples described above. For example, the steps recited in any method or process embodiment may be executed in any appropriate order and are not limited to the explicit order presented in the specific examples. Additionally, the components and/or elements recited in any system embodiment may be combined in a variety of permutations to produce substantially the same result as the present technology and are accordingly not limited to the specific configuration recited in the specific examples.

[0081] Benefits, other advantages and solutions to problems have been described above with regard to particular embodiments. Any benefit, advantage, solution to problems or any element that may cause any particular benefit, advantage or solution to occur or to become more pronounced, however, is not to be construed as a critical, required or essential feature or component. [0082] The terms "comprises", "comprising", or any variation thereof, are intended to reference a non-exclusive inclusion, such that a process, method, article, composition, system, or apparatus that comprises a list of elements does not include only those elements recited, but may also include other elements not expressly listed or inherent to such process, method, article, composition, system, or apparatus. Other combinations and/or modifications of the above-described structures, arrangements, applications, proportions, elements, materials or components used in the practice of the present technology , in addition to those not specifically recited, may be varied or otherwise particularly adapted to specific environments, manufacturing specifications, design parameters or other operating requirements without departing f om the general principles of the same.

[0083] The present technology has been described above with reference to an exemplary embodiment. However, changes and modifications may be made to the exemplary embodiment without departing from the scope of the present technology. These and other changes or modifications are intended to be included within the scope of the present technology.

Claims

CLAIMS hat is claimed is:

1. A method of treating a human glioblastoma tumor comprising glioma cells, comprising;

adding a therapeutical [y effective amount of the aurintricarboxylic acid to the glioma cells,

wherein the therapeutically effective amount of the aurintricarboxylic acid at least partial ly inhibits chemotactic migration of the glioma cells that exhibit elevated Fnl4 expression,

2. The method of claim 1, wherein the therapeutically effective amount of the aurintricarboxylic acid sensitizes the glioma cells to treatment with a chemotherapeutic agent,

3. The method of claim 2, wherein the chemotherapeutic agent comprises temozolomide.

4. The method of claim 1, wherein the therapeutically effective amount of the aurintricarboxylic acid sensitizes the glioma cells to treatment with radiation,

5. The method of claim 1, wherein the therapeutically effective amount of the aurintricarboxylic acid sensitizes the glioma cells to treatment with both a chemotherapeutic agent and radiation,

6. The method of claim 5, wherein the chemotherapeutic agent comprises temozolomide.

7. The method of claim 1, further comprising assaying the glioma cells for elevated Fill 4 expression as compared to noncancerous glial cells prior to adding the effective amount of the aurintricarboxylic acid.

8. A method of treating a human patient diagnosed with a cancer that exhibits elevated Fnl4 expression as compared to noncancer cel ls of the same cell type, comprising; administering a therapeutically effective amount of the aurin tricarboxylic acid to the patient,

wherein the therapeutically effective amount of the aurintricarboxylic acid sensitizes the cancer ceils to cell death from treatment with a chemotherapeutic agent; and

wherem the sensitization of the cancer cells is at least partially selective for targeting the cancer cells.

9. The method of claim 8, wherem the therapeutically effective amount is up to 50 mg/kg body wei ght of the patient.

10. The method of claim 8, wherein the cancer is a glioblastoma.

11. The method of claim 10, wherein the therapeutically effective amount of the aurintricarboxylic acid is administered to the patient by adding the aurintricarboxylic acid directly to the glioblastoma intracramally,

12. The method of claim 11, wherein the therapeutically effective amount of the aurintricarboxylic acid is in a concentration of approximately 10μΜ.

13. The method of claim 8 wherem the therapeutically effective amount of the aurintricarboxylic acid is administered to the patient intravenously.

14. The method of claim 8, wherein the therapeutically effective amount of the aurintricarboxylic acid further sensitizes the cancer cells to radiation therapy.

15. A method of treating cancer in a subject, the method comprising the steps of:

administering a pharmaceutical composition comprising a therapeutically effective amount of aurintricarboxylic acid (ATA) to the subject with cancer; and

administering a treatment modality that is known to treat the cancer,

16. The method of claim 15, wherein the treatment modality is selected from the group consisting of a chemotherapeutic agent and radiation therapy.

17. The method of claim 16, wherem the treatment modality comprises both a

chemotherapeutic agent and radiation therapy.

18. The method of claim 15, wherein the treatment modality comprises temozolomide.

19. The method of claim 15, wherein the cancer comprises glioblastoma.

20. The method of claim 19, wherein the glioblastoma is invasive.