Darryl T. Martin

Surface modified nanoparticles enhance transurothelial penetration and delivery of survivin siRNA in treating bladder cancer

Penetration of the bladder permeability barrier (BPB) is a major challenge when treating bladder diseases via intravesical delivery. To increase transurothelial migration and tissue and tumor cell uptake, poly(lactic-co-glycolic acid; PLGA) nanoparticles (NP) were modified by addition of a low molecular weight (2.5 or 20 kDa) positively charged mucoadhesive polysaccharide, chitosan, to the NP surface. In designing these NPs, we balanced the adhesive properties of chitosan with the release and bioactivity of the siRNA. Chitosan-functionalized NPs demonstrated increased binding to and uptake in intravesically instilled mouse bladders and human ureter at 10 times the level of unmodified NPs. Furthermore, we extended the bioactivity of survivin siRNA in vitro for up to 9 days and demonstrated a decrease in proliferation when using chitosan-modified NPs relative to unmodified NPs. In addition, treatment of xenograft tumors with chitosan-modified NPs that encapsulate survivin siRNA (NP-siSUR-CH2.5) resulted in a 65% reduction in tumor volume and a 75% decrease in survivin expression relative to tumors treated with blank chitosan NPs (NP-Bk-CH2.5). Our low molecular weight chitosan delivery system has the capacity to transport large amounts of siRNA across the urothelium and/or to the tumor site, thus increasing therapeutic response. Mol Cancer Ther; 13(1); 71–81. ©2013 AACR.

Nanoparticles for urothelium penetration and delivery of the histone deacetylase inhibitor belinostat for treatment of bladder cancer

UNLABELLED Nearly 40% of patients with non-invasive bladder cancer will progress to invasive disease despite locally-directed therapy. Overcoming the bladder permeability barrier (BPB) is a challenge for intravesical drug delivery. Using the fluorophore coumarin (C6), we synthesized C6-loaded poly(lactide-co-glycolide) (PLGA) nanoparticles (NPs), which were surface modified with a novel cell penetrating polymer, poly(guanidinium oxanorbornene) (PGON). Addition of PGON to the NP surface improved tissue penetration by 10-fold in intravesically-treated mouse bladder and ex vivo human ureter. In addition, NP-C6-PGON significantly enhanced intracellular uptake of NPs compared to NPs without PGON. To examine biological activity, we synthesized NPs that were loaded with the histone deacetylase (HDAC) inhibitor belinostat (NP-Bel-PGON). NP-Bel-PGON exhibited a significantly lower IC50 in cultured bladder cancer cells, and sustained hyperacetylation, when compared to unencapsulated belinostat. Xenograft tumors treated with NP-Bel-PGON showed a 70% reduction in volume, and a 2.5-fold higher intratumoral acetyl-H4, when compared to tumors treated with unloaded NP-PGON. FROM THE CLINICAL EDITOR These authors demonstrate that PLGA nanoparticles with PGON surface functionalization result in greatly enhanced cell penetrating capabilities, and present convincing data from a mouse model of bladder cancer for increased chemotherapy efficacy.

Blocking of the ATP sensitive potassium channel ameliorates the ischaemia‐reperfusion injury in the rat testis

There is increasing evidence that the effects of administered ATP sensitive potassium (KATP) channel openers or blockers during ischaemia are still controversial in many organs/tissues. Testicular torsion detorsion which causes ischaemia‐reperfusion (IR) injury, cannot be predicted, thus an effective drug should be administered during or after the ischaemia. The aim of this study was to examine whether the administration of KATP channel openers or blockers during ischaemia ameliorates IR injury in the testis. Eight‐week‐old male Sprague–Dawley rats were subjected to 2 h right testicular ischaemia followed by 24 h reperfusion. The selective mitochondrial (mito) KATP channel blocker, 5‐hydroxydecanoate (5‐HD) (40 mg/kg), the non‐selective KATP channel blocker glibenclamide (5 mg/kg), the selective mito KATP channel opener diazoxide (10 mg/kg) and the non‐selective KATP channel opener cromakalim (300 μg/kg) were administered intraperitoneally 15 min prior to the ischaemia or 75 min after the induction of ischaemia. Tissue damage was evaluated by malondialdehyde concentration, myeloperoxidase activity, histological evaluation and TdT‐mediated dUTP nick end labelling assay in the testis. There was a significant increase in oxidative stress, neutrophil infiltration, histological damage and apoptosis in the testicular IR model. A significant reduction in the testicular IR injury was observed with the administration of glibenclamide, but not 5‐HD, diazoxide or cromakalim during ischaemia. The administration of non‐selective KATP channel blocker glibenclamide ameliorated the testicular IR injury. On the other hand, the selective mito KATP channel blocker, 5‐HD and KATP channel openers did not reduce the testicular IR injury. These data suggest that blocking of the membrane KATP channel may have a protective effect during the testicular ischaemia. Glibenclamide could be an effective drug to manage the post‐ischaemic injury caused by the testicular torsion‐detorsion.

Controlled Cell Proliferation on an Electrochemically Engineered Collagen Scaffold

Therapies for corneal disease and injury often rely on artificial implants, but integrating cells into synthetic corneal materials remains a significant challenge. The electrochemically formed collagen-based matrix presented here is non-toxic to cells and controls the proliferation in the corneal fibroblasts seeded onto it. Histology and biomolecular studies show a behavior similar to corneal stromal cells in a native corneal environment. Not only is this result an important first step toward developing a more realistic, multi-component artificial cornea, but it also opens possibilities for using this matrix to control and contain the growth of cells in engineered tissues.

Olmesartan ameliorates urinary dysfunction in the spontaneously hypertensive rat via recovering bladder blood flow and decreasing oxidative stress.

As hypertension (HT) is one of the risk factors for lower urinary tract symptoms, we investigated the effect of an angiotensin II type I receptor blocker, olmesartan, on bladder dysfunction in the spontaneously hypertensive rat (SHR).

Effect of Silodosin, an Alpha1A-Adrenoceptor Antagonist, on Ventral Prostatic Hyperplasia in the Spontaneously Hypertensive Rat

Background A decreased prostatic blood flow could be one of the risk factors for benign prostatic hyperplasia/benign prostatic enlargement. The spontaneously hypertensive rat (SHR) shows a chronic prostatic ischemia and hyperplastic morphological abnormalities in the ventral prostate. The effect of silodosin, a selective alpha1A-adrenoceptor antagonist, was investigated in the SHR prostate as a prostatic hyperplasia model focusing on prostatic blood flow. Methods Twelve-week-old male SHRs were administered perorally with silodosin (100 μg/kg/day) or vehicle once daily for 6 weeks. Wistar Kyoto (WKY) rats were used as normotensive controls and were treated with the vehicle. The effect of silodosin on blood pressure and prostatic blood flow were estimated and then the prostates were removed and weighed. The tissue levels of malondialdehyde (MDA), interleukin-6 (IL-6), chemokine (C-X-C motif) ligand 1/cytokine-induced neutrophil chemoattractant 1 (CXCL1/CINC1), tumor necrosis factor-alpha (TNF-α), transforming growth factor beta 1 (TGF-β1), basic fibroblast growth factor (bFGF) and alpha-smooth muscle actin (α-SMA) were measured. The histological evaluation was also performed by hematoxylin and eosin staining. Results There was a significant increase in blood pressure, prostate weight, prostate body weight ratio (PBR), tissue levels of MDA, IL-6, CXCL1/CINC1, TNF-α, TGF-β1, bFGF and α-SMA in the SHR compared to the WKY rat. The ventral prostate in the SHR showed the morphological abnormalities compared to the WKY rat. Prostatic blood flow was decreased in the SHR. However, treatment with silodosin significantly restored the decreased prostatic blood flow in the SHR. Moreover, silodosin normalized tissue levels of MDA, IL-6, CXCL1/CINC1, TNF-α, TGF-β1, bFGF and α-SMA, and it ameliorated ventral prostatic hyperplasia in the SHR excluding blood pressure. Silodosin decreased PBR but not prostate weight in the SHR. Conclusions Silodosin can inhibit the progression of prostatic hyperplasia through a recovery of prostatic blood flow.

Protective effect of hydroxyfasudil, a Rho kinase inhibitor, on ventral prostatic hyperplasia in the spontaneously hypertensive rat.

Rho kinase (ROCK) pathway is associated with various cellular functions, such as smooth muscle contraction, inflammatory response, and cell proliferation. The spontaneously hypertensive rat (SHR) is commonly used genetically hypertensive rat model which develops hyperplastic morphological abnormalities in the ventral prostate. We investigated whether administration of hydroxyfasudil, a ROCK inhibitor, could reduce the levels of growth factors, inflammatory markers, and morphological abnormalities in the ventral prostate of the SHR.

Abstract B21: Chemoresistant bladder cancer cells induces epithelial mesenchymal transition

Although chemotherapeutic treatments are efficacious in a number of cancers, some cancer cells ultimately will become resistant to chemotherapy. It has been suggested that chronic chemotherapy-resistance is linked to the induction of epithelial mesenchymal transition (EMT). EMT is associated with increased tumor invasiveness, development of angiogenesis, and the development of drug resistance and is linked to more aggressive cancer. The objective of this study is to determine if a link exists between mitomycin C (MMC)-resistant RT-4 bladder cancer cells and the induction of EMT. EMT is a process where cells acquire molecular alterations characterized by loss of cell-cell adhesion, repression of E-cadherin expression, and increased cell mobility. Clinical studies have shown that as bladder cancer grade increases, tumors typically exhibit a decreased expression of E-cadherin, a cell-cell adhesion molecule. In addition, there is an increase in N-cadherin expression in muscle invasive bladder tumors. We have shown that MMC-resistant human RT-4 bladder cancer cells demonstrate a decrease in cell-cell adhesion, a higher invasion potential, and a higher CD44(+)/CD24(-) ratio (96.4%) compared to parental human RT-4 bladder cancer cells. In addition, the invasion capacity of MMC-resistant RT-4 bladder cancer cells was higher than that of parental cells. The MMC-resistant RT-4 bladder cancer cells expressed higher levels of N-cadherin and numerous apoptotic related proteins including pSTAT3, RAD17, Bcl-X and catalase. Using an exon array we showed that the MMC-resistant RT-4 bladder cancer cells up-regulate several EMT markers including vimentin (77 times), transforming growth factor -beta (27 times), and Zinc finger E-box-binding homeobox 1 (33 times) and down-regulate E-cadherin (71 times). In summary, in our MMC-resistant RT4 bladder cancer cells, EMT was activated, and there was an increase in apoptotic related proteins as well as an increase N-cadherin expression. Our data suggest a link between chemotherapy-resistant bladder cancer cells and EMT. Deactivating EMT and down-regulating apoptotic related proteins may offer a novel strategy for overcoming bladder cancer drug resistance. Citation Format: Jingchun Liu, Darryl T. Martin, Marcia A. Wheeler, Robert M. Weiss. Chemoresistant bladder cancer cells induces epithelial mesenchymal transition. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Drug Sensitivity and Resistance: Improving Cancer Therapy; Jun 18-21, 2014; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(4 Suppl): Abstract nr B21.