A number of pre-clinical studies using mouse and rat pain models have been completed. There are numerous rodent pre-clinical models that are commonly used in the industry in projecting the efficacy of various therapeutic agents.
Figure 1 depicts the anatomy of the pain nerve fibers. They can be divided into large myelinated and small thinly myelinated (Ad) or unmyelinated (C) fibers, often referred to as “small pain fibers.”
Peripheral nerve
Peripheral nerve is comprised of large mechano-sensitive (myelinated Ab) and small pain (Ad and C) fibers.
Figure 1 Peripheral nerve anatomy.
he most commonly used animal model is chronic construction injury (CCI). In this model, the rodent’s sciatic nerve is damaged and ligated back. This pain model usually persists for seven (7) weeks, which is sufficient to conduct pre-clinical studies on compounds of interest.
The CCI model produces unilateral peripheral mononeuropathy, and it has been observed that symptoms in this rat model correspond to causalgia or complex regional pain syndrome in patients. Figure 2 outlines the CCI model.
Figure 2 CCI model description.
Damage is conducted only on one side, while the second side is sham operation control (no injury). Thus, the same mouse or rat can serve as a negative control for pain (to compare how this specific mouse reacts with external stress in the presence of pain versus the absence of pain). Once the damage followed by ligation is complete, the same mice are treated or not treated, in our case with the anti-MMP-9 or anti-MMP-14 antibody, or with the negative control, irrelevant IgG antibody. The effects are compared.
Pain intensity is measured by two separate parameters:
1. Threshold of pressure in grams necessary for paw withdrawal from a mechanical stress agent
2. Frequency paw withdrawal within a two-minute period from a specific stress agent
Figure 3 is an example of the data generated with our therapeutic antibodies in the CCI model.
Figure 3 demonstrates the efficacy of the anti-MMP-9 antibody in the pain model. The panel on the left shows a significant reduction in the pressure threshold for paw withdrawal, thus signifying significant pain in the affected paw. When treated with the anti-MMP-9 antibody, there is significant restoration of the initial, pre-injury pressure threshold.A similar pattern is observed in the frequency of paw withdrawal in the panel on the right. After the introduction of injury, the frequency of withdrawal is elevated from zero to approximately 90%. After treating the mice with Releviate Therapeutics’ anti-MMP antibody, the frequency of withdrawal goes from 90% to 30%, while treatment with the control irrelevant antibody does not produce any significant effect. This data demonstrates that our therapeutic antibody effectively and specifically relieves pain.
Figure 4 demonstrates the efficacy of the anti-MMP-9 antibody. Panels a through f represent separate animal model experiments using the anti-MMP-9 antibody. The conclusion from each experiment is on the left of the figure.
Spinal MMP-9 is Required and Sufficient to Induce Neuropathic Pain
Figure 4 Efficacy of anti-MMP-9 human antibody in the rat spinal nerve ligation (SNL) model.
(a) Persistent IT infusion of MMP-9 inhibitor delays mechanical allodynia in SNL rats.
(b) Reversal of mechanical allodynia by TIMP-1, endogenous MMP-9 inhibitor, in SNL rats
(c) Pretreatment of MMP-9 siRNA (2 x 5 µg, i.t.) delays mechanical allodynia in SNL rats
(d) IT MMP-9 induces reversible mechanical allodynia in rats
(e) MMP-9 null mice show a reduction in SNL-induced spontaneous pain at early times
(f) MMP-9 null mice show a reduction of mechanical allodynia at early times after SNL (* p<0.05, compared to wild-type mice, n=6).(a)
Figure 5 – Route of Administration Graph
In 2021, Releviate completed the study to select the best and most efficacious, according to the relevant animal studies, route of administration, where intrathecal, intravenous, and intra-dorsal-root-ganglion administration have been tested. Figure 5 summarizes the meaningful efficacy outcomes of the study.
It is clear from the data that RLVT-9-03 significantly relieves pain in the PSNL model. More importantly, it is persistent action, as compared to two-hour efficacy of the current standard of care, such as gabapentin. It persists beyond Day 21, if IV administration is used. Another very important conclusion of this study (drawn from the efficacy of the RLVT-9-03 in the IV route of administration) is that, in the case of neuropathic pain, the therapeutic, disease-modifying, agent does not have to cross the blood-brain barrier. This is not surprising since the main morphological difference of the neural fibers affected by the neuropathy is that the fibers become permeable and thus highly sensitized. The main advantage of our therapy is the ability to reverse such morphology, restoring normal fiber integrity and sensitivity.
Summary of pre-clinical data
MMP-14/MT-1-MMP
• Reverses the established neuropathic pain (mechanical allodynia)
• Reduces neuropathology (axon demyelination and degeneration)
• Reduces peripheral and central neuroinflammation, and maladaptive neuroplasticity
• Anti-MMP14 antibody displayed very high specificity, thus reducing the risk of side effects
Anti-MMP-9 human antibody (RLVT-9-03)
• An injury-specific target (no effect on normal nociception)
• Prevents and reduces neuropathic pain development. Exhibits sustained (weeks) efficacy by IV delivery
• Reduces neuropathology, peripheral and central neuroinflammation and maladaptive neuroplasticity and protects neurons
• Very high specificity and potency
• Efficacy sustainable for the duration of the model
For more information on the pre-clinical data and results of the pre-clinical studies, please refer to the PUBLICATION LIST.