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.

Smoking is a major risk factor for urothelial cell carcinoma of the bladder (UCC). Mutations in the FGFR3 and TP53 genes have been shown to define two distinct pathways in superficial papillary and invasive UCC disease, respectively. We investigated the relationship between smoking and these mutations by means of denaturing high performance liquid chromatography and sequencing for 110 primary UCC of the bladder. This study included 48 current smokers, 31 ex-smokers and 31 non-smokers. Thirty-five of the tumors were stage pTa, 40 pT1 and 35 > or =pT2. Fourteen of the tumors were grade 1, 37 were grade 2 and 59 grade 3. Smoking was associated with high stage (P = 0.03) and high grade tumors (P = 0.006). Twenty-two of the 110 tumors studied harbored TP53 mutations (20%) and 43 harbored FGFR3 mutations (39%). Odds ratios (OR) were higher for TP53 mutations in current smokers [OR, 2.25; 95% confidence interval (95% CI), 0.65-7.75] and ex-smokers (OR, 1.62; 95% CI, 0.41-6.42) than in non-smokers. Double TP53 mutations and the A:T-->G:C TP53 mutation pattern was found only in current smokers. Patients with the FGFR3(wild-type)/TP53(mutated) genotype had significantly higher levels of tobacco consumption, as measured in pack-years (P = 0.01). Smoking influenced neither the frequency nor the pattern of FGFR3 mutations. Our results suggest that smoking is associated with invasive and high grade UCCs, at initial presentation, and influenced TP53 or the molecular pathway defined by these mutations. In contrast, FGFR3 mutations are not affected by smoking and probably result from endogenous alterations. These data have potential implications for clinical management and prevention strategies.

What ' s known on the subject? and What does the study add? Evidence has emerged identifying folic acid supplementation as a potential risk factor for cancer development or progression. Long-term folic acid supplementation has been shown to increase the risk of prostate cancer development by three-fold. Sarcosine is a byproduct of folate metabolism and has been proposed as a biomarker for aggressive prostate cancer phenotypes.We looked at the effects of physiologically relevant levels of folic acid on in vitro prostate cancer cell growth and invasion, and demonstrated that higher levels can have the effect of increasing both of these biological processes. We also show that these changes toward a more aggressive phenotype are not linked to increased sarcosine levels, however other metabolic pathways may be involved. OBJECTIVES• To investigate the effects of different folic acid concentrations on the growth and invasiveness of prostate cancer cell lines.• To determine if observed changes are correlated with changes in levels of the potential prostate cancer biomarker, sarcosine, a byproduct of folate metabolism. MATERIALS AND METHODS• The prostate cancer cell lines PC-3, LNCaP and DU145 were cultured in media containing 4, 20 or 100 n M of folic acid and assayed for growth over 9 days by counting viable cells at 3-day intervals, or for invasion by passage through a Matrigel-coated transwell membrane.• Cells grown in the different folic acid media were collected and subjected to metabolomic analysis by gas chromatography and mass spectrometry to measure levels of intracellular sarcosine. RESULTS• The results show that higher levels of folic acid can increase cell growth in PC-3 and LNCaP prostate cancer cell lines, and may also increase the invasive capacity of PC-3, LNCaP and DU145 cells.• We did not observe a correlation between increased invasion from higher folic acid concentrations and levels of sarcosine, but there were signifi cant changes in other metabolites in cells grown in higher levels of folic acid. CONCLUSION• These fi ndings suggest that folic acid has an important and potentially negative role in prostate cancer progression. KEYWORDprostate cancer , folic acid , invasion , metabolomics , sarcosine