The tumor cell-derived hyaluronidase HYAL-1 degrades hyaluronic acid (HA) into pro-angiogenic fragments that support tumor progression. Although HYAL-1 is a critical determinant of tumor progression and a marker for cancer diagnosis and metastasis prediction, it has not been evaluated as a target for cancer therapy. Similarly, sulfated hyaluronic acid (sHA) has not been evaluated for biological activity, although it is a HAase inhibitor. In this study we show that sHA is a potent inhibitor of prostate cancer. sHA blocked the proliferation, motility and invasion of LNCaP, LNCaP-AI, DU145 and LAPC-4 prostate cancer cells, also inducing caspase 8-dependent apoptosis associated with downregulation of Bcl-2 and phospho-Bad. sHA inhibited Akt signaling including androgen receptor (AR) phosphorylation, AR-activity, NFkb activation and VEGF expression. These effects were traced to a blockade in complex formation between PI3K and HA receptors and to a transcriptional downregulation of HA receptors, CD44 and RHAMM, along with PI3K inhibition. Angiogenic HA fragments or overexpression of myristoylated-Akt or HA receptors blunted these effects of sHA, implicating a feedback loop between HA receptors and PI3K/Akt signaling in the mechanism of action. In an animal model, sHA strongly inhibited LNCaP-AI prostate tumor growth without causing weight loss or apparent serum-organ toxicity. Inhibition of tumor growth was accompanied by a significant decrease in tumor angiogenesis and an increase in apoptosis index. Taken together, our findings offer mechanistic insights into the tumor-associated HA-HAase system and a preclinical proof-of-concept of the safety and efficacy of sHA to control prostate cancer growth and progression.

FANCA is a component of the Fanconi anemia (FA) core complex that activates DNA interstrand crosslink repair by monoubiquitination of FANCD2. Here, we report that purified FANCA protein catalyzes bidirectional single-strand annealing (SA) and strand exchange (SE) at a level comparable to RAD52, while a disease-causing FANCA mutant, F1263Δ, is defective in both activities. FANCG, which directly interacts with FANCA, dramatically stimulates its SA and SE activities. Alternatively, FANCB, which does not directly interact with FANCA, does not stimulate this activity. Importantly, five other patient-derived FANCA mutants also exhibit deficient SA and SE, suggesting that the biochemical activities of FANCA are relevant to the etiology of FA. A cell-based DNA double-strand break (DSB) repair assay demonstrates that FANCA plays a direct role in the single-strand annealing sub-pathway (SSA) of DSB repair by catalyzing SA, and this role is independent of the canonical FA pathway and RAD52.

Background: 3Ј repair exonuclease 1 (TREX1) is a DNase involved in autoimmune disorders and the antiviral response. Results: TREX1 also degrades single-stranded RNA or RNA in a RNA/DNA hybrid molecule. Conclusion: TREX1 is a human homolog of Escherichia coli RNase T. Significance: The novel RNase activity of TREX1 is crucial for understanding its physiological role.

PURPOSE Current treatments for metastatic RCC (mRCC) do not extend survival beyond a few months. Sorafenib (SF) is a targeted drug approved for mRCC, but it has modest efficacy. Hymecromone is a nontoxic dietary supplement with some antitumor activity at high doses (450 – 3000 mg/day). HC inhibits hyaluronic acid (HA) synthesis. HA promotes tumor growth and metastasis. We recently showed that HA-receptors CD44 and RHAMM are potential predictors of mRCC. We examined the anti-tumor properties of HC, SF, and their combination in RCC models. METHODS Using proliferation, clonogenic and apoptosis assays, effects of HC (0–32 μg/ml), SF (0–3.2 μg/ml) and HC+SF were examined in RCC cells (Caki–1, 786–O, ACHN, A498) and endothelial cells (HMVEC–L, HUVEC). Boyden chamber was used for motility and invasion assays. Apoptosis indicators, HA receptors, EGFR and c-Met were evaluated by immunoblotting. Efficacy of HC, SF and HC+SF was evaluated in the SF-resistant Caki–1 xenograft model. RESULTS HC+SF synergistically inhibited proliferation (>95%), motility/invasion (65%) and capillary formation (76%) in RCC and/or endothelial cells, and induced apoptosis by 8-fold (P<0.001). HC+SF inhibited HA synthesis and HA addition reversed the cytotoxicity of HC+SF. HC+SF up-regulated pro-apoptotic indicators and downregulated Mcl-1, CD44, RHAMM, phospho-EGFR and phospho-cMet levels. In all assays, HC and SF alone were ineffective. Oral administration of HC (50–200mg/kg) plus SF (30mg/kg) eradicated Caki–1 tumor growth without toxicity. HC and SF alone were ineffective. CONCLUSION This is the first study that demonstrates combination of SF with HC a non–toxic dietary supplement is highly effective in controlling RCC.

MUS81-EME1 is a DNA endonuclease involved in replication-coupled repair of DNA interstrand cross-links (ICLs). A prevalent hypothetical role of MUS81-EME1 in ICL repair is to unhook the damage by incising the leading strand at the 3′ side of an ICL lesion. In this study, we report that purified MUS81-EME1 incises DNA at the 5′ side of a psoralen ICL residing in fork structures. Intriguingly, ICL repair protein, Fanconi anemia complementation group A protein (FANCA), greatly enhances MUS81-EME1-mediated ICL incision. On the contrary, FANCA exhibits a two-phase incision regulation when DNA is undamaged or the damage affects only one DNA strand. Studies using truncated FANCA proteins indicate that both the N- and C-moieties of the protein are required for the incision regulation. Using laser-induced psoralen ICL formation in cells, we find that FANCA interacts with and recruits MUS81 to ICL lesions. This report clarifies the incision specificity of MUS81-EME1 on ICL damage and establishes that FANCA regulates the incision activity of MUS81-EME1 in a damage-dependent manner.

Mutations in the SETX gene, which encodes Senataxin, are associated with the progressive neurodegenerative diseases ataxia with oculomotor apraxia 2 (AOA2) and amyotrophic lateral sclerosis 4 (ALS4). To identify the causal defect in AOA2, patient-derived cells and SETX knockouts (human and mouse) were analyzed using integrated genomic and transcriptomic approaches. A genome-wide increase in chromosome instability (gains and losses) within genes and at chromosome fragile sites was observed, resulting in changes to gene-expression profiles. Transcription stress near promoters correlated with high GCskew and the accumulation of R-loops at promoter-proximal regions, which localized with chromosomal regions where gains and losses were observed. In the absence of Senataxin, the Cockayne syndrome protein CSB was required for the recruitment of the transcription-coupled repair endonucleases (XPG and XPF) and RAD52 recombination protein to target and resolve transcription bubbles containing R-loops, leading to genomic instability. These results show that transcription stress is an important contributor to SETX mutation-associated chromosome fragility and AOA2.

Background CXCR4 and CXCR7 are seven transmembrane chemokine receptors of the stroma-derived factor (SDF-1). CXCR4, but not CXCR7, has been examined in bladder cancer (BCa). We examined the functional and clinical significance of CXCR7 in BCa. Methods CXCR4 and CXCR7 levels were measured in BCa cell lines, tissues (normal = 25; BCa = 44) and urine specimens (n=186) by quantitative PCR and/or immunohistochemistry. CXCR7 function in BCa cells were examined by transient transfections using a CXCR7 expression vector or siRNA. Results In BCa cell lines CXCR7 mRNA levels were 5–37-fold higher than CXCR4 levels. Transient overexpression of CXCR7 in BCa cell lines promoted growth and chemotactic motility. CXCR7 co-localized and formed a functional complex with EGF-receptor, PI3-kinase/Akt, Erk and src and induced their phosphorylation. CXCR7 also induced upregulation of cyclin-D1 and bcl-2. Suppression of CXCR7 expression reversed these effects and induced apoptosis. CXCR7 mRNA levels and CXCR7 staining scores were significantly (5–10-fold) higher in BCa tissues than in normal tissues (P<0.001). CXCR7 expression independently associated with metastasis (P=0.019) and disease specific mortality (P=0.03). CXCR7 was highly expressed in endothelial cells in high-grade BCa tissues when compare to low-grade BCa and normal bladder. CXCR7 levels were elevated in exfoliated urothelial cells from high-grade BCa patients (P=0.0001; 90% sensitivity; 75% specificity); CXCR4 levels were unaltered. Conclusion CXCR7 promotes BCa cell proliferation and motility plausibly through EGF-receptor and Akt-signaling. CXCR7 expression is elevated in BCa tissues and exfoliated cells and is associated with high-grade and metastasis.

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