The SPOP-ITCH Signaling Axis Protects Against Prostate Cancer Metastasis.

Prostate cancer is one of the most common causes of cancer incidence and death in men, with the mortality caused primarily by the late-stage and metastatic forms of the disease. The mechanisms and molecular markers for prostate cancer metastasis are not fully understood. Speckle type Poz Protein (SPOP) is an E3 ubiquitin ligase adaptor that is often mutated in prostate cancer. In this study, we sequenced the SPOP gene in 198 prostate cancer patients and found 16 mutations in the cohort. Multivariate analysis revealed that SPOP mutations correlated with the clinical stage of the disease and strongly with metastasis. We identified ITCH as a candidate protein for SPOP-mediated degradation via mass spectrometry. We demonstrated the interaction between SPOP and ITCH, and found that the SPOP F133L mutation disrupted the SPOP-ITCH interaction, leading to a subsequent increase in the ITCH protein level. Further, we found that the SPOP knockdown led to higher levels of Epithelial- mesenchymal transition (EMT) proteins and increased cell invasion. Together, our results highlight the functional significance of the SPOP-ITCH pathway in prostate cancer metastasis.

KAT5 promotes invasion and metastasis through C-MYC stabilization in ATC.

Anaplastic thyroid cancer (ATC) is an aggressive cancer with poor clinical prognosis. However, mechanisms driving ATC aggressiveness is not well known. Components of the DNA damage response (DDR) are frequently found mutated or aberrantly expressed in ATC. The goal of this study is to establish the functional link between histone acetyltransferase lysine (K) acetyltransferase 5 (KAT5, a critical DDR protein) and ATC invasiveness using clinical, in vitro and in vivo models. We analyzed the expression of KAT5 by immunohistochemistry and assessed its relationship with metastasis and overall survival in 82 ATC patients. Using cellular models, we established functional connection of KAT5 expression and C-MYC stabilization. We then studied the impact of genetically modified KAT5 expression on ATC metastasis in nude mice. In clinical samples, there is a strong correlation of KAT5 expression with ATC metastasis (P = 0.0009) and overall survival (P = 0.0017). At the cellular level, upregulation of KAT5 significantly promotes thyroid cancer cell proliferation and invasion. We also find that KAT5 enhances the C-MYC protein level by inhibiting ubiquitin-mediated degradation. Further evidence reveals that KAT5 acetylates and stabilizes C-MYC. Finally, we prove that altered KAT5 expression influences ATC lung metastases in vivo. KAT5 promotes ATC invasion and metastases through stabilization of C-MYC, demonstrating it as a new biomarker and therapeutic target for ATC.

Functional roles of Speckle-Type Poz (SPOP) Protein in Genomic stability.

Understanding the functional significance of the essential elements in maintaining genomic stability provides insights into the process of tumor initiation and progression, and predicts therapeutic responses. One such element that has recently attracted significant attention is the Speckle-Type Poz Protein (SPOP), an E3 ubiquitin ligase adaptor protein. SPOP is frequently mutated or has altered expression in various cancers, including prostate, renal and endometrial. SPOP is involved in the regulation of proteasome-mediated degradation of several oncoproteins. Moreover, recent data also indicate SPOP's direct involvement in the DNA damage response. SPOP mutants induce alternations in the DNA damage repair pathway by promoting the error-prone Non-homologous end joining (NHEJ) pathway. SPOP has been linked with significant functions in cellular signaling pathways and cancer suppression. This mini-review will discuss recent findings regarding SPOP's role in genomic stability in the pathological setting.

Addition of carbonic anhydrase 9 inhibitor SLC-0111 to temozolomide treatment delays glioblastoma growth in vivo.

Tumor microenvironments can promote stem cell maintenance, tumor growth, and therapeutic resistance, findings linked by the tumor-initiating cell hypothesis. Standard of care for glioblastoma (GBM) includes temozolomide chemotherapy, which is not curative, due, in part, to residual therapy-resistant brain tumor-initiating cells (BTICs). Temozolomide efficacy may be increased by targeting carbonic anhydrase 9 (CA9), a hypoxia-responsive gene important for maintaining the altered pH gradient of tumor cells. Using patient-derived GBM xenograft cells, we explored whether CA9 and CA12 inhibitor SLC-0111 could decrease GBM growth in combination with temozolomide or influence percentages of BTICs after chemotherapy. In multiple GBMs, SLC-0111 used concurrently with temozolomide reduced cell growth and induced cell cycle arrest via DNA damage in vitro. In addition, this treatment shifted tumor metabolism to a suppressed bioenergetic state in vivo. SLC-0111 also inhibited the enrichment of BTICs after temozolomide treatment determined via CD133 expression and neurosphere formation capacity. GBM xenografts treated with SLC-0111 in combination with temozolomide regressed significantly, and this effect was greater than that of temozolomide or SLC-0111 alone. We determined that SLC-0111 improves the efficacy of temozolomide to extend survival of GBM-bearing mice and should be explored as a treatment strategy in combination with current standard of care.

Cks1 proteasomal degradation is induced by inhibiting Hsp90-mediated chaperoning in cancer cells.

PURPOSE: Cks1, a conformationally heterogenous 9 kDa protein, is markedly overexpressed in cancer cells and contributes to tumor development. Cks1 is an essential component of the SCF-Skp2 ubiquitin ligase complex that targets the Cdk inhibitors p27(Kip1) and p21(Cip1). Cks1 is known to interact with the Hsp90-Cdc37 chaperone machinery, although whether this facilitates its conformational maturation and stability is not known. To test whether abrogating the chaperone function of Hsp90 could destabilize Cks1, we examined the effects of treating different cancer cell lines with the benzoquinone ansamycin 17-allylamino geldanamycin (17-AAG), a compound that selectively binds Hsp90 and potently inhibits its ATP-dependent chaperone activity. METHODS: The effect of Hsp90 inhibition using 17-AAG on Cks1 protein and associated cell cycle proteins including Skp2, p27(Kip1), p21(Cip1), and Cdk1 in cancer cells was determined by Western blotting. Ubiquitination analysis was carried out by transfecting cells with an HA-ubiquitin plasmid and specifically immunoprecipitating Cks1 to examine polyubiquitinated species. Flow cytometry was utilized to examine the effects of Hsp90 inhibition on cell cycle profiles. RESULTS: Here, we demonstrate for the first time that inhibition of Hsp90 utilizing 17-AAG destabilizes Cks1 in cancer cells by promoting its ubiquitination and proteasomal degradation. 17-AAG-induced Cks1 depletion was accompanied by concomitant decreases in Skp2 and Cdk1. 17-AAG treatment also induced G2/M accumulation in MCF-7 breast carcinoma cells, and G1 accumulation in the colon carcinoma lines HCT116 and SW620. CONCLUSIONS: We conclude that perturbing the Hsp90 pathway could provide a useful therapeutic strategy in tumors driven by Cks1 overexpression.