The use of light as a means of therapy for bladder cancer (BC) has a long history but has been hampered by a lack of tumor specificity and therefore, damage to the normal bladder mucosa. Here, we describe a targeted form of photo-therapy called photoimmunotherapy (PIT) which targets EGFR-expressing BC. Anti-EGFR antibody panitumumab (pan) was labeled with the photo-absorber (PA), IRDye 700Dx (IR700), to create a pan IR700 antibody-PA conjugate which is activated by near-infrared radiation (NIR). BC tissue microarray (TMA) and BC cell lines were analyzed for expression of EGFR. Mechanism of PIT induced cell death was studied using proliferation assays, transmission electron microscopy (TEM), and production of reactive oxygen species. Finally, the in vivo effect was studied in xenografts. EGFR staining of TMAs showed that while most BCs have expression of EGFR to a varying degree, squamous cell carcinomas (SCC) have the highest expression of EGFR. Pan IR700 activated by NIR light rapidly killed UMUC-5 cells, a bladder SCC line. Pan alone, pan IR700 without NIR, or NIR alone had no effect on cells. TEM demonstrated that cell death is due to necrosis. Singlet oxygen species contributed towards cell death. NIR-PIT with pan IR700 reduced growth compared to only pan IR700 treated UMUC-5 xenograft tumors. PIT is a new targeted treatment for bladder cancer. Pan IR700-induced PIT selectively kills EGFR-expressing BC cells in vitro and in vivo and therefore warrants further therapeutic studies in orthotopic xenografts of BC and ultimately in patients.

Bladder cancer (BC) is heterogeneous and expresses various cell surface targets. Photoimmunotherapy (PIT) involves monoclonal antibodies (MAbs) conjugated to a photoabsorber (PA), IR Dye 700Dx, and then activated by near infra-red light (NIR) to specifically target tumors. We have demonstrated that tumors expressing EGFR can be targeted with PIT. However, PIT may be less effective when a tumor lacks “overwhelming” expression of a single target such as EGFR. We present a combinatorial PIT approach for targeting BC expressing EGFR and HER2, using PA- labeled panitumumab (pan) and trastuzumab (tra), respectively. Human BC tissues and cell lines were analyzed for EGFR and HER2 expression. Efficacy of PA-labeled MAbs singly and in combination was analyzed. About 45% of BC tissues stain for both EGFR and HER2. In vitro, the combination of pan IR700 and tra IR700 with NIR was more efficacious than either agent alone. Tumor xenografts treated with combination PIT showed significant tumor growth retardation. Combination PIT is a promising approach for treating BC with low/moderate expression of surface receptors. In addition, given the molecular heterogeneity of bladder cancer, targeting more than one surface receptor may allow for more effective cell death across different bladder tumors.

Heat shock protein 90 (HSP90) inhibition is an attractive strategy for cancer treatment. Several HSP90 inhibitors have shown promising effects in clinical oncology trials. However, little is known about HSP90 inhibition-mediated bladder cancer therapy. Here, we report a quantitative proteomic study that evaluates alterations in protein expression and histone post-translational modifications (PTMs) in bladder carcinoma in response to HSP90 inhibition. We show that 5 HSP90 inhibitors (AUY922, ganetespib, SNX2112, AT13387, and CUDC305) potently inhibited the proliferation of bladder cancer 5637 cells in a dose- and time-dependent manner. Our proteomic study quantified 518 twofold up-regulated and 811 twofold down-regulated proteins common to both AUY922 and ganetespib treatment. Bioinformatic analyses revealed that those differentially expressed proteins were involved in multiple cellular processes and enzyme-regulated signaling pathways, including chromatin modifications and cell death-associated pathways. Furthermore, quantitative proteome studies identified 14 types of PTMs with 93 marks on the core histones, including 34 novel histone marks of butyrylation, citrullination, 2-hydroxyisobutyrylation, methylation, O-GlcNAcylation, propionylation, and succinylation in AUY922- and ganetespib-treated 5637 cells. Together, this study outlines the association between proteomic changes and histone PTMs in response to HSP90 inhibitor treatment in bladder carcinoma cells, and thus intensifies the understanding of HSP90 inhibition-mediated bladder cancer therapeutics.

Cycling is associated with many health benefits, but also with the risk of injury. Trends in bicycle-related injuries are difficult to assess because the majority of nonfatal injuries sustained while cycling are not reported to police and thus are not included in traffic statistics. 1 We sought to evaluate trends in adult cycling injuries and hospital admissions in the United States using emergency department data.Methods | The National Electronic Injury Surveillance System (NEISS) is a national probability sample of approximately 100 emergency departments that gathers product-related injury data. 2 We queried the NEISS for injuries associated with bicycles (codes 5033 and 5040) from 1998 to 2013. The University of California, San Francisco, institutional review board gave the study exempt status.The number of bicycle-related injuries in adults aged 18 years or older was recorded in 2-year intervals. We used the NEISS complex sample design to calculate population projections of cycling-related injuries, which were then divided by US Census data to produce incidence per 100 000 persons. Adjustment for age was performed using the direct method.

Perineal nodular induration (PNI), or biker's nodule, is a rare, bothersome, pseudotumour. Herein, we describe the surgical technique used to treat a healthy cyclist who developed an enlarging PNI for five years that grew into a perineal mass. The mass prevented him from cycling due to worsening discomfort and heaviness. The PNI-associated mass was successfully removed by wide surgical excision and a local advancement flap. Subsequently, the patient resumed cycling. Histopathology report demonstrated a benign lesion with abundant ropy collagen with native smooth muscle, vessels, and rare fibroblast-like spindle cells. With the increasing popularity of cycling, PNI may become more common, and health providers should be aware of this rare entity and how it can be safely removed.

The protein kinase D (PKD) family of proteins are important regulators of tumor growth, development, and progression. CRT0066101, an inhibitor of PKD, has antitumor activity in multiple types of carcinomas. However, the effect and mechanism of CRT0066101 in bladder cancer are not understood. In the present study, we show that CRT0066101 suppressed the proliferation and migration of four bladder cancer cell lines in vitro. We also demonstrate that CRT0066101 blocked tumor growth in a mouse flank xenograft model of bladder cancer. To further assess the role of PKD in bladder carcinoma, we examined the three PKD isoforms and found that PKD2 was highly expressed in eight bladder cancer cell lines and in urothelial carcinoma tissues from the TCGA database, and that short hairpin RNA (shRNA)-mediated knockdown of PKD2 dramatically reduced bladder cancer growth and invasion in vitro and in vivo, suggesting that the effect of the compound in bladder cancer is mediated through inhibition of PKD2. This notion was corroborated by demonstrating that the levels of phospho-PKD2 were markedly decreased in CRT0066101-treated bladder tumor explants. Furthermore, our cell cycle analysis by flow cytometry revealed that CRT0066101 treatment or PKD2 silencing arrested bladder cancer cells at the G2/M phase, the arrest being accompanied by decreases in the levels of cyclin B1, CDK1 and phospho-CDK1 (Thr161) and increases in the levels of p27 and phospho-CDK1 (Thr14/Tyr15). Moreover, CRT0066101 downregulated the expression of Cdc25C, which dephosphorylates/activates CDK1, but enhanced the activity of the checkpoint kinase Chk1, which inhibits CDK1 by phosphorylating/inactivating Cdc25C. Finally, CRT0066101 was found to elevate the levels of Myt1, Wee1, phospho-Cdc25C (Ser216), Gadd45α, and 14-3-3 proteins, all of which reduce the CDK1-cyclin B1 complex activity. These novel findings suggest that CRT0066101 suppresses bladder cancer growth by inhibiting PKD2 through induction of G2/M cell cycle arrest, leading to the blockade of cell cycle progression.

Purpose: Kidney stone prevention relies on the 24-hour urine collection to diagnose metabolic abnormalities and direct dietary and pharmacological therapy. While its use is guideline supported for high risk and interested patients, evidence that the test can accurately predict recurrence or treatment response is limited. We sought to critically reassess the role of the 24-hour urine collection in stone prevention. Materials and Methods: In addition to a MEDLINEÒ search to identify controlled studies of dietary and pharmacological interventions, evidence supporting the AUA (American Urological Association) and EAU (European Association of Urology) guidelines for metabolic stone prevention were evaluated. Additionally, the placebo arms of these studies were examined to assess the stone clinic effect, that is the impact of regular office visits without specific treatment on stone recurrence. Results: The 24-hour urine test has several limitations, including the complexity of interpretation, the need for repeat collections, the inability to predict stone recurrence with individual parameters and supersaturation values, the unclear rationale of laboratory cutoff values and the difficulty of determining collection adequacy. Only 1 prospective trial has compared selective dietary recommendations based on 24-hour urine collection results vs general dietary instructions. While the trial supported the intervention arm, significant limitations to the study were found. Placebo arms of intervention trials have noted a 0% to 61% decrease in stone recurrence rate and a remission rate during the study of 20% to 86%. Conclusions: Whether all recurrent stone formers benefit from 24-hour urine collection has not been established. Additional comparative effectiveness trials are needed to determine which stone former benefits from selective therapy, as guided by the 24-hour urine collection.

Purpose Previous studies have shown that ischemia alters gene expression in normal and malignant tissues. There are no studies that evaluated effects of ischemia in renal tumors. This study examines the impact of ischemia and tissue procurement conditions on RNA integrity and gene expression in renal cell carcinoma. Experimental Design Ten renal tumors were resected without renal hilar clamping from 10 patients with renal clear cell carcinoma. Immediately after tumor resection, a piece of tumor was snap frozen. Remaining tumor samples were stored at 4C, 22C and 37C and frozen at 5, 30, 60, 120, and 240 minutes. Histopathologic evaluation was performed on all tissue samples, and only those with greater than 80% tumor were selected for further analysis. RNA integrity was confirmed by electropherograms and quantitated using RIN index. Altered gene expression was assessed by paired, two-sample t-test between the zero time point and aliquots from various conditions obtained from the same tumor. Results One hundred and forty microarrays were performed. Some RNA degradation was observed 240 mins after resection at 37C. The expression of over 4,000 genes was significantly altered by ischemia times or storage conditions. The greatest gene expression changes were observed with longer ischemia time and warmer tissue procurement conditions. Conclusion RNA from kidney cancer remains intact for up to 4 hours post surgical resection regardless of storage conditions. Despite excellent RNA preservation, time after resection and procurement conditions significantly influence gene expression profiles. Meticulous attention to pre-acquisition variables is of paramount importance for accurate tumor profiling.