Liposome Based Intravesical Therapy Targeting Nerve Growth Factor Ameliorates Bladder Hypersensitivity in Rats with Experimental Colitis.

Pelvic organ cross sensitization is considered to contribute to overlapping symptoms in chronic pelvic pain syndrome. Nerve growth factor over expression in the bladder is reportedly involved in the symptom development of bladder pain syndrome/interstitial cystitis. We examined whether a reduction of over expressed nerve growth factor in the bladder by intravesical treatment with liposome and oligonucleotide conjugates would ameliorate bladder hypersensitivity in a rat colitis model. Adult female rats were divided into 1) a control group, 2) a colitis-oligonucleotide group with intracolonic TNBS (2,4,6-trinitrobenzene sulfonic acid) enema and intravesical liposome-oligonucleotide treatments, 2) a colitis-saline group with intracolonic TNBS and intravesical saline treatments, 4) a sham oligonucleotide group with intravesical liposome-oligonucleotide treatment without colitis and 5) a sham-saline group with intravesical saline treatment without colitis. Liposomes conjugated with nerve growth factor antisense oligonucleotide or saline solution were instilled in the bladder and 24 hours later colitis was induced by TNBS enema. Effects of nerve growth factor antisense treatment were evaluated by pain behavior, cystometry, molecular analyses and immunohistochemistry 10 days after TNBS treatment. In colitis-oligonucleotide rats nerve growth factor antisense treatment ameliorated pain behavior and decreased a reduction in the intercontraction interval in response to acetic acid stimulation as well as nerve growth factor expression in the bladder mucosa. All were enhanced in colitis-saline rats compared to sham rats. Nerve growth factor over expression in the bladder mucosa and bladder hypersensitivity induced after colitis were decreased by intravesical application of liposome-oligonucleotide targeting nerve growth factor. This suggests that local antinerve growth factor therapy could be effective treatment of bladder symptoms in chronic pelvic pain syndrome.

Kawamorita N ，Yoshikawa S ，Kashyap M ，Tyagi P ，Arai Y ，Chancellor MB ，Yoshimura N ... -  《-》

Effect of Intravesical Liposome-Based Nerve Growth Factor Antisense Therapy on Bladder Overactivity and Nociception in a Rat Model of Cystitis Induced by Hydrogen Peroxide.

Majima T ，Tyagi P ，Dogishi K ，Kashyap M ，Funahashi Y ，Gotoh M ，Chancellor MB ，Yoshimura N ... -  《-》

Down-regulation of nerve growth factor expression in the bladder by antisense oligonucleotides as new treatment for overactive bladder.

Nerve growth factor over expression in the bladder has a role in overactive bladder symptoms via the mediation of functional changes in bladder afferent pathways. We studied whether blocking nerve growth factor over expression in bladder urothelium by a sequence specific gene silencing mechanism would suppress bladder overactivity and chemokine expression induced by acetic acid. Female Sprague-Dawley® rats anesthetized with isoflurane were instilled with 0.5 ml saline, scrambled or TYE™ 563 labeled antisense oligonucleotide targeting nerve growth factor (12 μM) alone or complexed with cationic liposomes for 30 minutes. The efficacy of nerve growth factor antisense treatments for acetic acid induced bladder overactivity was assessed by cystometry. Bladder nerve growth factor expression levels and cellular distribution were quantified by immunofluorescence staining and enzyme-linked immunosorbent assay. Effects on bladder chemokine expression were measured by Luminex® xMAP® analysis. Liposomes were needed for bladder uptake of oligonucleotide, as seen by the absence of bright red TYE 563 fluorescence in rats instilled with oligonucleotide alone. At 24 hours after liposome-oligonucleotide treatment baseline bladder activity during saline infusion was indistinct in the sham and antisense treated groups with a mean ± SEM intercontraction interval of 348 ± 55 and 390 ± 120 seconds, respectively. Acetic acid induced bladder overactivity was shown by a decrease in the intercontraction interval to a mean of 33.2% ± 4.0% of baseline in sham treated rats. However, the reduction was blunted to a mean of 75.8% ± 3.4% of baseline in rats treated with liposomal antisense oligonucleotide (p <0.05). Acetic acid induced increased nerve growth factor in the urothelium of sham treated rats, which was decreased by antisense treatment, as shown by enzyme-linked immunosorbent assay and reduced nerve growth factor immunoreactivity in the urothelium. Increased nerve growth factor in bladder tissue was associated with sICAM-1, sE-selectin, CXCL-10 and 1, leptin, MCP-1 and vascular endothelial growth factor over expression, which was significantly decreased by nerve growth factor antisense treatment (p <0.01). Acetic acid induced bladder overactivity is associated with nerve growth factor over expression in the urothelium and with chemokine up-regulation. Treatment with liposomal antisense suppresses bladder overactivity, and nerve growth factor and chemokine expression. Local suppression of nerve growth factor in the bladder could be an attractive approach for overactive bladder. It would avoid the systemic side effects that may be associated with nonspecific blockade of nerve growth factor expression.

Kashyap M ，Kawamorita N ，Tyagi V ，Sugino Y ，Chancellor M ，Yoshimura N ，Tyagi P ... -  《-》

A Prospective, Multicenter, Double-Blind, Randomized Trial of Bladder Instillation of Liposome Formulation OnabotulinumtoxinA for Interstitial Cystitis/Bladder Pain Syndrome.

Intravesical instillation of liposomal formulated botulinum toxin A (lipotoxin) has shown therapeutic effects as treatment of refractory overactive bladder without needle injections. We assessed lipotoxin to treat refractory interstitial cystitis/bladder pain syndrome. This 2-center, double-blind, randomized, placebo controlled, physician initiated study enrolled patients with refractory interstitial cystitis/bladder pain syndrome. A total of 31 patients were assigned to intravesical instillation of lipotoxin (onabotulinumtoxinA 200 U with 80 mg sphingomyelin), 28 were assigned to onabotulinumtoxinA 200 U in normal saline and 31 were assigned to normal saline alone. The primary end point was the average change in O'Leary-Sant symptom scores, including ICSI (Interstitial Cystitis Symptom Index) and ICPI (Interstitial Cystitis Problem Index) between baseline and 4 weeks after treatment. Other end points included the average changes in a 3-day voiding diary, a visual analog scale for pain and a global response assessment of patient satisfaction. Improvements in the pain scale and O'Leary-Sant symptom scores occurred in all 3 groups by 4 weeks after treatment. Lipotoxin instillation was associated with a statistically significant decrease in O'Leary-Sant symptom scores (mean ± SD 7.38 ± 8.75), ICSI (4.00 ± 4.28), ICPI (3.35 ± 5.11) and the visual analog scale pain scale (1.64 ± 2.52), and an increase in the global response assessment (1.35 ± 1.28). However, there was no difference in improvement among the 3 groups. No significant adverse events were found in any group. Lipotoxin failed to demonstrate a positive proof of concept compared to onabotulinumtoxinA or placebo. However, a single intravesical instillation of lipotoxin was associated with decreased interstitial cystitis/bladder pain syndrome symptoms compared to baseline in patients with moderate to severe interstitial cystitis/bladder pain syndrome. The effect was likely due to a significant placebo effect.

Chuang YC ，Kuo HC 《-》

Therapeutic effects of nerve growth factor-targeting therapy on bladder overactivity in rats with prostatic inflammation.

The present study examined the effect of liposomes conjugated with antisense oligonucleotide of nerve growth factor (NGF-OND) on local overexpression of NGF and bladder overactivity using rats with prostatic inflammation (PI). Male Sprague-Dawley rats were divided into three groups: (1) Control group; intact rats, (2) PI-NS group; rats with PI and intravesical instillation of normal saline (NS), (3) PI-OND group; rats with PI and intravesical instillation of NGF-OND. On Day 0, PI was induced by intraprostatic 5%-formalin injection. On Day 14, NGF-OND or NS was instilled directly into the bladder after laparotomy. On Day 28, therapeutic effects of NGF-OND were evaluated by awake cystometry and histological analysis as well as reverse-transcription polymerase chain reaction measurements of messenger RNA (mRNA) levels of NGF in the bladder and prostate, inflammatory markers in the prostate, C-fiber afferent markers, and an A-type K+ channel α-subunit (Kv 1.4) in L6-S1 dorsal root ganglia (DRG). Intravesical NFG-OND treatment reduced PI-induced overexpression of NGF in both bladder and prostate, and reduced PI-induced bladder overactivity evident as longer intercontraction intervals in association with reductions of TRPV1 and TRPA1 mRNA expression levels in DRG. mRNA expression of Kv1.4 in DRG was reduced after PI, but improved in the PI-OND group. These results indicate that NGF locally expressed in the bladder is an important mediator inducing bladder overactivity with upregulation of C-fiber afferent markers and downregulation of an A-type K+ channel subunit in DRG following PI, and that liposome-based, local NGF-targeting therapy could be effective for not only bladder overactivity and afferent sensitization, but also PI. Thus, local blockade of NGF in the bladder could be a therapeutic modality for male LUTS due to BPH with PI.

Igarashi T ，Tyagi P ，Mizoguchi S ，Saito T ，Furuta A ，Suzuki Y ，Egawa S ，Wang Z ，Yoshimura N ... -  《-》