Abstract4-Methylumbelliferone (4-MU) is a hyaluronic acid (HA) synthesis inhibitor with anticancer properties; the mechanism of its anticancer effects is unknown. We evaluated the effects of 4-MU on prostate cancer cells. 4-MU inhibited proliferation, motility, and invasion of DU145, PC3-ML, LNCaP, C4-2B, and/or LAPC-4 cells. At IC 50 for HA synthesis (0.4 mmol/L), 4-MU induced >3-fold apoptosis in prostate cancer cells, which could be prevented by the addition of HA. 4-MU induced caspase-8, caspase-9, and caspase-3 activation, PARP cleavage, upregulation of Fas-L, Fas, FADD and DR4, and downregulation of bcl-2, phosphorylated bad, bcl-XL, phosphorylated Akt, phosphorylated IKB, phosphorylated ErbB2, and phosphorylated epidermal growth factor receptor. At IC 50 , 4-MU also caused >90% inhibition of NF-κB reporter activity, which was prevented partially by the addition of HA. With the exception of caveolin-1, HA reversed the 4-MU-induced downregulation of HA receptors (CD44 and RHAMM), matrix-degrading enzymes (MMP-2 and MMP-9), interleukin-8, and chemokine receptors (CXCR1, CXCR4, and CXCR7) at the protein and mRNA levels. Expression of myristoylated-Akt rescued 4-MU-induced apoptosis and inhibition of cell growth and interleukin-8, RHAMM, HAS2, CD44, and MMP-9 expression. Oral administration of 4-MU significantly decreased PC3-ML tumor growth (>3-fold) when treatment was started either on the day of tumor cell injection or after the tumors became palpable, without organ toxicity, changes in serum chemistry, or body weight. Tumors from 4-MU-treated animals showed reduced microvessel density (∼3-fold) and HA expression but increased terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cells and expression of apoptosisrelated molecules. Therefore, the anticancer effects of 4-MU, an orally bioavailable and relatively nontoxic agent, are primarily mediated by inhibition of HA signaling. Cancer Res; 70(7); 2613-23. ©2010 AACR.

BACKGROUND: Cancer biomarkers are the backbone for the implementation of individualized approaches to bladder cancer (BCa). Hyaluronic acid (HA) and all 7 members of the HA family, that is, HA synthases (HA1, HA2, HA3), HYAL-1 hyaluronidase, and HA receptors (CD44s, CD44v, and RHAMM), function in tumor growth and progression. However, the diagnostic and prognostic potential of these 7 HA family members has not been compared simultaneously in any cancer. We evaluated the diagnostic and prognostic potential of HA family members in BCa. METHODS: Using quantitative PCR and immunohistochemistry, expression of HA family members was evaluated in prospectively collected bladder tissues (n ¼ 72); mean and median follow-up were 29.6 AE 5.3 and 24 months, respectively. Transcript levels were also measured in exfoliated urothelial cells from urine specimens (n ¼ 148). RESULTS: Among the HA family members, transcript levels of the HA synthases, HYAL-1, CD44v, and RHAMM were 4-to 16-fold higher in BCa tissues than in normal tissues (P < .0001); however, CD44s levels were lower. In univariate and multivariate analyses, tumor stage (P ¼ .003), lymph node invasion (P ¼ .033), HYAL-1 (P ¼ .019), and HAS1 (P ¼ .027) transcript levels, and HYAL-1 staining (P ¼ .021) were independently associated with metastasis. Tumor stage (P ¼ .019) and HYAL-1 (P ¼ .046) transcript levels were also associated with disease-specific mortality. Although HA synthase and HYAL-1 transcript levels were elevated in exfoliated urothelial cells from BCa patients, the combined HAS2-HYAL-1 expression detected BCa with an overall sensitivity of 85.4% and a specificity of 79.5% and predicted BCa recurrence within 6 months (P ¼ .004; RR ¼ 6.7). CONCLUSIONS: HYAL-1 and HAS1 expression predicted BCa metastasis, and HYAL-1 expression also predicted disease-specific survival. Furthermore, the combined HAS2-HYAL-1 biomarker detected BCa and significantly predicted its recurrence.

4-MU is an effective nontoxic, oral chemopreventive, and therapeutic agent that targets PCa development, growth, and metastasis by abrogating HA signaling.

Benign prostatic hyperplasia (BPH) is one of the most common conditions affecting men. BPH can lead to a number of symptoms for patients commonly referred to as lower urinary tract symptoms (LUTS).Over the last decade, increased modifiable risk factors, such as metabolic disease and obesity, have resulted in an increased incidence of BPH. This increasing incidence has brought about a multitude of treatment modalities in the last two decades. With so many treatment modalities available, physicians are tasked with selecting the optimal therapy for their patients. Current therapies can first be divided into medical or surgical intervention. Medical therapy for BPH includes 5-alpha-reductase inhibitors and alpha-blockers, or a combination of both. Surgical interventions include a conventional transurethral resection of the prostate (TURP), as well as newer modalities such as bipolar TURP, holmium laser enucleation of the prostate (HoLEP), Greenlight and thulium laser, and prostatic urethral lift (PUL). Emerging therapies in this field must also be further investigated for safety and efficacy. This narrative review attempts to consolidate current and emerging treatment options for BPH and highlights the need for additional investigation on optimizing treatment selection.

Background:Molecular markers of clinical outcome may aid in designing targeted treatments for bladder cancer. However, only a few bladder cancer biomarkers have been examined as therapeutic targets.Methods:Data from The Cancer Genome Atlas (TCGA) and bladder specimens were evaluated to determine the biomarker potential of the hyaluronic acid (HA) family of molecules – HA synthases, HA receptors and hyaluronidase. The therapeutic efficacy of 4-methylumbelliferone (4MU), a HA synthesis inhibitor, was evaluated in vitro and in xenograft models.Results:In clinical specimens and TCGA data sets, HA synthases and hyaluronidase-1 levels significantly predicted metastasis and poor survival. 4-Methylumbelliferone inhibited proliferation and motility/invasion and induced apoptosis in bladder cancer cells. Oral administration of 4MU both prevented and inhibited tumour growth, without dose-related toxicity. Effects of 4MU were mediated through the inhibition of CD44/RHAMM and phosphatidylinositol 3-kinase/AKT axis, and of epithelial–mesenchymal transition determinants. These were attenuated by HA, suggesting that 4MU targets oncogenic HA signalling. In tumour specimens and the TCGA data set, HA family expression correlated positively with β-catenin, Twist and Snail expression, but negatively with E-cadherin expression.Conclusions:This study demonstrates that the HA family can be exploited for developing a biomarker-driven, targeted treatment for bladder cancer, and 4MU, a non-toxic oral HA synthesis inhibitor, is one such candidate.

This is the first study that showed a decreasing gradient of Gal-3 expression in benign, adjacent-benign and tumor tissues. Gal-3 expression may be useful in predicting biochemical recurrence.

Purpose: Poor prognosis of patients with muscle-invasive bladder cancer (BC) that often metastasizes drives the need for discovery of molecular determinants of BC progression.Chondroitin sulfate proteoglycans, including CD44, regulate cancer progression; however, the identity of a chondroitinase (Chase) that cleaves chondroitin sulfate from proteoglycans is unknown. HYAL-4 is an understudied gene suspected to encode a Chase, with no known biological function. We evaluated HYAL-4 expression and its role in BC. Experimental design:In clinical specimens HYAL-4 wild-type (Wt) and V1 expression was evaluated by RT-qPCR, immunohistochemistry and/or immunoblotting; a novel assay measured Chase activity. Wt and V1 were stably expressed or silenced in normal urothelial and three BC cell lines. Transfectants were analyzed for stem cell phenotype, invasive signature and tumorigenesis, and metastasis in four xenograft models, including orthotopic bladder.Results: HYAL-4 expression, specifically a novel splice variant (V1), was elevated in bladder tumors; Wt expression was barely detectable. V1 encoded a truncated 349 amino acid protein that was secreted. In BC tissues, V1 levels associated with metastasis and cancer-specificsurvival with high efficacy and encoded Chase activity. V1 cleaved chondroitin-6-sulfate from CD44, increasing CD44 secretion. V1 induced stem cell phenotype, motility/invasion, and an invasive signature. CD44 knockdown abrogated these phenotypes. V1-expressing urothelial cells developed angiogenic, muscle-invasive tumors. V1-expressing BC cells formed tumors at low-density and formed metastatic bladder tumors when implanted orthotopically. Conclusion:Our study discovered the first naturally-occurring eukaryotic/human Chase and connected it to disease pathology, specifically cancer. V1-Chase is a driver of malignant BC and potential predictor of outcome in BC patients.

What's known on the subject? and What does the study add? A significant proportion of patients diagnosed with prostate cancer do not require immediate treatment and could be managed by active surveillance, which usually includes serial measurements of prostate‐specific antigen (PSA) levels and regular biopsies. The rate of rise in PSA levels, which could be calculated as PSA velocity or PSA doubling time, was previously suggested to be associated with the biological aggressiveness of prostate cancer. Although these parameters are obvious candidates for predicting tumour progression in active surveillance patients, earlier studies that examined this topic provided conflicting results. Our analysis showed that PSA velocity and PSA doubling time calculated at different time‐points, by different methods, over different intervals, and in different sub‐groups of active surveillance patients provide little if any prognostic information. Although we found some significant associations between PSA velocity and the risk of progression as determined by biopsy, the actual clinical significance of this association was small. Furthermore, PSA velocity did not add to the predictive accuracy of total PSA. Objective To study whether prostate‐specific antigen (PSA) velocity (PSAV) and PSA doubling time (PSADT) are associated with biopsy progression in patients managed by active surveillance. Patients and Methods Our inclusion criteria for active surveillance are biopsy Gleason sum <7, two or fewer positive biopsy cores, ≤20% tumour present in any core, and clinical stage T1–T2a. Changes in any of these parameters during the follow‐up that went beyond these limits are considered to be progression. This study included 250 patients who had at least one surveillance biopsy, an available PSA measured no earlier than 3 months before diagnosis, and at least one PSA measurement before each surveillance biopsy. We evaluated the association between PSA kinetics and progression at successive surveillance biopsies in different sub‐groups of patients by calculating the area under the curve (AUC) as well as sensitivity and specificity of different thresholds. Results Over a median follow‐up of 3.0 years, the disease of 64 (26%) patients progressed. PSADT was not associated with biopsy progression, whereas PSAV was only weakly associated with progression in certain sub‐groups. However, incorporation of PSAV in models including total PSA resulted in a moderate increase in AUC only when the entire cohort was analysed. In other sub‐groups the predictive accuracy of total PSA was not significantly improved by adding PSAV. Conclusions Our findings confirm that PSA kinetics should not be used in decision‐making in patients with low‐risk prostate cancer managed by active surveillance. Regular surveillance biopsies should remain as the principal method of monitoring cancer progression in these men.