Functional Signature Ontology-Based Identification and Validation of Novel Therapeutic Targets and Natural Products for the Treatment of Cancer

Multiple studies have revealed that Ras-driven tumors acquire vulnerabilities by adapting cellular mechanisms that promote uncontrolled proliferation and suppress apoptosis. Kinase Suppressor of Ras 1 (KSR1) modulates ERK activation downstream of oncogenic Ras, and knockdown of KSR1 selectively kills malignant, Ras-driven cancer cells, but does not kill immortalized, non-transformed human colon epithelial cells (HCECs). KSR1-/- mice are fertile and phenotypically normal, but resistant to Ras-driven tumor formation suggesting KSR1 represents a vulnerability in cancer cells. To identify additional vulnerabilities in cancer, a screening approach termed Functional Signature Ontology (FUSION) was used to screen 14,355 genes and 1,200 natural product fractions in K-Ras-mutant HCT116 colon cancer cells for functional similarity to KSR1 and a selective requirement in colon cancer cells. FUSION identified numerous targets including TIMELESS, WDR5, and an AMPK inhibitor, 5’-hydroxy-staurosporine. Downstream of oncogenic signaling, TIMELESS is constitutively overexpressed in multiple types of cancer and required for increased cancer cell proliferation. TIMELESS depletion increases γH2AX, a marker of DNA damage, and triggers downstream G2/M arrest via increased CHK1 and CDK1 phosphorylation. Wee1 or CHK1 inhibition in combination with TIMELESS depletion demonstrates at least additive effects suggesting this combination may be efficacious for the treatment of cancer. WDR5 is overexpressed, and WDR5 depletion reduces cell viability in colon cancer cells by reducing H3K4Me3 and increasing γH2AX, which further sensitizes cells to radiation-induced DNA damage. WDR5 inhibition also reduces colon cancer cell viability, but less so than WDR5 depletion. The catalytic, kinase-containing a2 subunit isoform of AMPK is expressed at variable levels in colon cancer cells and is selectively required for colon cancer cell survival suggesting that AMPK kinase inhibition may be a useful component of cancer therapeutic strategies. FUSION identified 5´-hydroxy-staurosporine as a competitive inhibitor of AMPK that is selectively toxic to colon cancer cells. Our results demonstrate the ability of FUSION to reveal functional similarities between genes, identify novel inhibitors, and expose oncogene-induced changes in cancer that promote proliferation and survival, but may also leave cancer cells vulnerable to targeted therapies

Rewards for Rights Ratification? Testing for Tangible and Intangible Benefits of Human Rights Treaty Ratification

Among the explanations for state ratification of human rights treaties, few are more common and widely accepted than the conjecture that states are rewarded for ratification by other states. These rewards are expected to come in the form of tangible benefits—foreign aid, trade, and investment—and intangible benefits such as praise, acceptance, and legitimacy. Surprisingly, these explanations for ratification have never been tested empirically. We summarize and clarify the theoretical underpinnings of “reward-for-ratification” theories and test these propositions empirically by looking for increased international aid, economic agreements, and public praise and recognition following ratification of four prominent human rights treaties. We find almost no evidence that states can expect increased tangible or intangible rewards after ratification. Given the lack of empirical support, alternative explanations seem more appealing for understanding human rights treaty ratification

Rewards for Rights Ratification? Testing for Tangible and Intangible Benefits of Human Rights Treaty Ratification

Among the explanations for state ratification of human rights treaties, few are more common and widely accepted than the conjecture that states are rewarded for ratification by other states. These rewards are expected to come in the form of tangible benefits—foreign aid, trade, and investment—and intangible benefits such as praise, acceptance, and legitimacy. Surprisingly, these explanations for ratification have never been tested empirically. We summarize and clarify the theoretical underpinnings of “reward-for-ratification” theories and test these propositions empirically by looking for increased international aid, economic agreements, and public praise and recognition following ratification of four prominent human rights treaties. We find almost no evidence that states can expect increased tangible or intangible rewards after ratification. Given the lack of empirical support, alternative explanations seem more appealing for understanding human rights treaty ratification

Radiation Therapy Improves Local Control in Juvenile Nasopharyngeal Angiofibroma following Disease Progression after Embolization and Surgical Resection: A Case Report

Juvenile nasopharyngeal angiofibroma (JNA) is a relatively uncommon, benign neoplasm of the nasopharynx that can be very difficult to diagnose early due to inconspicuous and seemingly harmless presenting symptoms. Early diagnosis and treatment of JNA are essential for a good prognosis. JNA typically responds well to radiation therapy (RT), but when it does not, the most appropriate next course of action has not been readily defined due to the limited occurrence and experience with this neoplasm. Herein, we describe a JNA patient, who continued to progress after surgery and 36 Gy of adjuvant radiation, but after an additional 14.4 Gy, he has remained in remission for over 2 years. An 11-year-old boy who presented with JNA underwent treatment with embolization and surgical resection. Unfortunately, the tumor progressed within 2 months of surgical intervention and he required RT for adequate local control. While undergoing RT, he again demonstrated signs of progression; so his radiation regimen was increased from 3,600 cGy in 20 fractions to 5,040 cGy in 28 fractions. Since completing RT, the tumor has continued to decrease in size, and the patient is stable and has been without signs of disease progression for over 24 months now. Thus, escalating the radiation regimen to 5,040 cGy may improve local control in rapidly progressive JNA

Highly Aggressive and Radiation-Resistant, “Atypical” and Silent Pituitary Corticotrophic Carcinoma: A Case Report and Review of the Literature

Background: Pituitary tumors typically remain silent unless interaction with nearby structures occurs. Rare subsets of pituitary tumors display aggressive phenotypes: highly mitotic, locally invasive, metastatic, chemotherapy and radiation resistant, etc. Disease progression and response to therapy is ill-defined in these subtypes, and their true prognostic potential is debated. Thus, identifying tumor characteristics with prognostic value and efficacious treatment options remains a challenge in aggressive pituitary tumors. Case Presentation: A 45-year-old female presented with a nonfunctioning corticotropic pituitary macroadenoma with biomarkers suggestive of an “atypical” subtype: Ki-67 of 8–12%, increased mitosis, and locally invasive. Despite resections and radiation, growth continued, eventually affecting her vision. Although histologically ACTH positive, the patient remained clinically asymptomatic. Twelve months later, an episode of Cushing’s disease-induced psychosis prompted a PET-CT scan, identifying sites of metastasis. Temozolomide was added to her medical regimen, and her metastatic liver lesions and boney metastases were treated with radiofrequency ablation and stereotactic body radiation therapy, respectively. Systemic treatment resulted in a drop in her ACTH levels, with her most recent scans/labs at 12 months following RFA suggesting remission. Conclusions: This is a unique presentation of a pituitary tumor, displaying characteristics of both clinically silent corticotropic and “atypical” macroadenoma subtypes. Although initially ACTH positive while clinically silent, the patient’s disease ultimately recurred metastatically with manifestations of Cushing’s disease and psychosis. With the addition of temozolomide to her treatment plan, her primary and metastatic sites have responded favorably to radiation therapy. Thus, the addition of temozolomide may be beneficial in the treatment of aggressive pituitary tumors

A Functional Signature Ontology (FUSION) screen detects an AMPK inhibitor with selective toxicity toward human colon tumor cells

AMPK is a serine threonine kinase composed of a heterotrimer of a catalytic, kinase-containing α and regulatory β and γ subunits. Here we show that individual AMPK subunit expression and requirement for survival varies across colon cancer cell lines. While AMPKα1 expression is relatively consistent across colon cancer cell lines, AMPKα1 depletion does not induce cell death. Conversely, AMPKα2 is expressed at variable levels in colon cancer cells. In high expressing SW480 and moderate expressing HCT116 colon cancer cells, siRNA-mediated depletion induces cell death. These data suggest that AMPK kinase inhibition may be a useful component of future therapeutic strategies. We used Functional Signature Ontology (FUSION) to screen a natural product library to identify compounds that were inhibitors of AMPK to test its potential for detecting small molecules with preferential toxicity toward human colon tumor cells. FUSION identified 5′-hydroxy-staurosporine, which competitively inhibits AMPK. Human colon cancer cell lines are notably more sensitive to 5′-hydroxy-staurosporine than are non-transformed human colon epithelial cells. This study serves as proof-of-concept for unbiased FUSION-based detection of small molecule inhibitors of therapeutic targets and highlights its potential to identify novel compounds for cancer therapy development

A New Milky Way Dwarf Galaxy in Ursa Major

In this Letter, we report the discovery of a new dwarf satellite to the Milky Way, located at ($\alpha_{2000}, \delta_{2000}$) $=$ (158.72,51.92) in the constellation of Ursa Major. This object was detected as an overdensity of red, resolved stars in Sloan Digital Sky Survey data. The color-magnitude diagram of the Ursa Major dwarf looks remarkably similar to that of Sextans, the lowest surface brightness Milky Way companion known, but with approximately an order of magnitude fewer stars. Deeper follow-up imaging confirms this object has an old and metal-poor stellar population and is $\sim$ 100 kpc away. We roughly estimate M$_V =$ -6.75 and $r_{1/2} =$ 250 pc for this dwarf. Its luminosity is several times fainter than the faintest known Milky Way dwarf. However, its physical size is typical for dSphs. Even though its absolute magnitude and size are presently quite uncertain, Ursa Major is likely the lowest luminosity and lowest surface brightness galaxy yet known.Comment: Replaced with ApJL accepted version. Includes some additional details, corrected references, and minor changes to Figure

Characterization of CDK(5) Inhibitor, 20-223 (aka CP668863) for Colorectal Cancer Therapy

Colorectal cancer (CRC) remains one of the leading causes of cancer related deaths in the United States. Currently, there are limited therapeutic options for patients suffering from CRC, none of which focus on the cell signaling mechanisms controlled by the popular kinase family, cyclin dependent kinases (CDKs). Here we evaluate a Pfizer developed compound, CP668863, that inhibits cyclin-dependent kinase 5 (CDK5) in neurodegenerative disorders. CDK5 has been implicated in a number of cancers, most recently as an oncogene in colorectal cancers. Our lab synthesized and characterized CP668863 – now called 20-223. In our established colorectal cancer xenograft model, 20-223 reduced tumor growth and tumor weight indicating its value as a potential anti-CRC agent. We subjected 20-223 to a series of cell-free and cell-based studies to understand the mechanism of its anti-tumor effects. In our hands, in vitro 20-223 is most potent against CDK2 and CDK5. The clinically used CDK inhibitor AT7519 and 20-223 share the aminopyrazole core and we used it to benchmark the 20-223 potency. In CDK5 and CDK2 kinase assays, 20-223 was ~3.5-fold and ~65.3-fold more potent than known clinically used CDK inhibitor, AT7519, respectively. Cell-based studies examining phosphorylation of downstream substrates revealed 20-223 inhibits the kinase activity of CDK5 and CDK2 in multiple CRC cell lines. Consistent with CDK5 inhibition, 20-223 inhibited migration of CRC cells in a wound-healing assay. Profiling a panel of CRC cell lines for growth inhibitory effects showed that 20-223 has nanomolar potency across multiple CRC cell lines and was on an average \u3e2-fold more potent than AT7519. Cell cycle analyses in CRC cells revealed that 20-223 phenocopied the effects associated with AT7519. Collectively, these findings suggest that 20-223 exerts anti-tumor effects against CRC by targeting CDK 2/5 and inducing cell cycle arrest. Our studies also indicate that 20-223 is a suitable lead compound for colorectal cancer therapy