Several weeks post-spinal cord injury (SCI) whatever the cause i.e. medical or traumatic, neurogenic detrusor overactivity (NDO) and detrusor sphincter dyssynergia (DSD) occur in most patients. These permanent lower urinary tract dysfunctions are responsible for the development of high bladder pressures and uncontrolled urinary incontinence because of spontaneous involuntary detrusor contractions. 

The pathophysiology of this lower urinary tract dysfunction relies on the development of an abnormal reflex at the spinal cord level. This abnormal spinal reflex is mostly due to neuroplasticity impairing bladder afferent C-fibers properties. Indeed, the C- fibers are normally silent during bladder filling and micturition. Peripheral and central release of neurotrophic factors post-SCI is responsible for a change in C-fibers activity. C-fibers become mechano-sensitive and drive an abnormal spinal micturition reflex.
The standard of care for NDO in SCI patients relies on pharmacological inhibition of bladder contractions to alleviate urinary incontinence with oral antimuscarinics or beta-3-agonists as 1st line and intra-detrusor botulinum toxin A (BoNT/A) injections under cystoscopy as 2nd line. In case of resistance or intolerance to antimuscarinics or beta-3-agonist +/- alpha blocker, imipramine is needed. BoNT/A injections must be repeated approximately every 9 months. These drugs inhibit the transmission of the neural signal between the parasympathetic neuron terminals and the detrusor muscle. Next to the need to repeat these injections about every 9 months there is also the risk of urinary retention in patients who can still void spontaneously. 

EG110A is a novel non-replicative recombinant HSV-1-derived vector that expresses the light chain of the botulinum toxin F (BoNT/F-LC) driven by the human calcitonin gene-related protein (hCGRP) promoter to achieve local sensory neuron-selective transgenic expression. The mechanism of action of the EG110A is based on the natural ability of HSV-1 to selectively infect peripheral neurons, especially afferent ones, and to express the BoNT/F-LC inside the targeted neurons leading to the cleavage of the VAMP2 protein that is essential for neurotransmission. The hCGRP promoter intends to reinforce the selectivity of the vectors for  C-fiber bladder afferents. The aim for EG110A is to provide a long-term efficacy, possibly lasting several years, for the treatment of NDO while preserving the ability of the bladder to contract and not cause urinary retention.

Here we investigated the ability of EG110A to inhibit bladder sensory C fibers in an acute non-clinical intravesical capsaicin model. Capsaicin acts on sensory nerves via vanilloid receptors. Acute intravesical capsaicin is a suitable model to assess afferent hyperactivity mimicking the pathophysiology of NDO and other functional urology conditions such as OAB (overactive bladder).

EG110A was injected in the detrusor muscle of adult female rats (2E+08 PFU in 40µL per rat) after open laparotomy exposing the bladder. Control animals received 40µL of vehicle buffer. One week after injection five rats injected with EG110A were euthanized and L6/S1 dorsal root ganglia (DRGs) were collected for the measurement of the transgene expression by ddPCR. At five weeks after injection, rats (n=14 rats per group) were anesthetized and a catheter connected to a pressure transducer was inserted in the bladder through the dome. The bladder was continuously perfused with saline during a stabilization period of 90 min followed by perfusion with 30 µM of capsaicin for 60 min. Intravesical pressure was recorded via cystometry throughout the experiment and parameters of reflex-evoked bladder contractions were measured. Data were computed over 15 min intervals during the capsaicin irritation period and normalized for each rat by the value of the stabilization period. Three control rats and two EG110A-injected rats could not be analyzed due to leakage of the bladder during cystometry experiment. Following cystometry measurements, L6/S1 DRGs were collected for RNA expression analysis. Expression of BoNT/F-LC was measured by digital droplet PCR (ddPCR) on cDNAs following reverse transcription.

ddPCR on DRGs showed a significant number of copies of BoNT/F-LC transcripts at one week following intradetrusor injection in all rats (Figure 1). BoNT/F-LC expression levels were not significantly different at 1 and 5 weeks after injection. No expression was seen in vehicle-injected rats. Body weight of rats increased similarly between vehicle- and EG110A-injected rats during the 5 weeks between intradetrusor injection and capsaicin irritation (data not shown). 5 weeks after injection, parameters measured during cystometry were not significantly changed during the stabilization period (data not shown). Intravesical capsaicin infusion triggered a rapid decrease of the interval between two voiding contractions (intercontraction interval, ICI) in vehicle treated rats (reduced to 36% of stabilization period). This decrease remained stable during the 60 min of capsaicin perfusion. The reduction of ICI was significantly inhibited in EG110A-treated animals (Figure 2) during the whole capsaicin irritation period. Infused volume between two voiding contractions, baseline and maximal pressure, voided volume and compliance were also significantly improved by EG110A upon capsaicin irritation. Post-void residual volumes measured at the end of cystometry were similar between vehicle and EG110A-treated groups.

Following local administration of EG110A in the detrusor, the vector EG110A is able to transduce afferent fibers to achieve a stable expression of BoNT/F-LC for at least 5 weeks in L6/S1 DRG sensory neurons. EG110A administration did not induce mortality nor deleterious effect on the body weight evolution during the 5 weeks of the experiment. Moreover, under normal condition with saline perfusion during the stabilization period, EG110A did not induce a significant change on cystometric parameters versus vehicle group nor did alter the voiding efficiency under saline or capsaicin perfusion. These results point it out the safety of the intradetrusor administration of EG110A. The main finding of this study is that during capsaicin perfusion, EG110A apparently counteracted bladder irritation elicited by capsaicin-mediated activation of TRPV1 receptors in C-fiber bladder afferents, as evidenced by a significant decrease in the frequency of micturition and increase in the bladder capacity. Finally, due to its selective effect on C-fibers, this study shows that EG110A improves ICI under capsaicin irritation without any change in normal bladder function. In addition, EG110A does not lead to urinary retention.

HSV1-based non replicative vectors are being developed as an efficient tool to achieve long term expression of a transgene in sensory neurons. EG110A is a promising approach to treat C-fiber related NDO and lower urinary tract disorders. EG110A has the potential to provide long term efficacy without adverse effects on voiding. Finally, these results show for the first time that intraneuronal expression of BoNT/F-LC can have a significant therapeutic potential.

Continence 7S1 (2023) 100817
DOI: 10.1016/j.cont.2023.100817