Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
Randomized Controlled Trial
. 2014 May 1;9(5):e96408.
doi: 10.1371/journal.pone.0096408. eCollection 2014.

Efficacy of a metalloproteinase inhibitor in spinal cord injured dogs

Jonathan M Levine  1 , Noah D Cohen  2 , Michael Heller  3 , Virginia R Fajt  4 , Gwendolyn J Levine  5 , Sharon C Kerwin  1 , Alpa A Trivedi  6 , Thomas M Fandel  6 , Zena Werb  7 , Augusta Modestino  3 , Linda J Noble-Haeusslein  8
Affiliations
Randomized Controlled Trial

Efficacy of a metalloproteinase inhibitor in spinal cord injured dogs

Jonathan M Levine et al. PLoS One. .

Abstract

Matrix metalloproteinase-9 is elevated within the acutely injured murine spinal cord and blockade of this early proteolytic activity with GM6001, a broad-spectrum matrix metalloproteinase inhibitor, results in improved recovery after spinal cord injury. As matrix metalloproteinase-9 is likewise acutely elevated in dogs with naturally occurring spinal cord injuries, we evaluated efficacy of GM6001 solubilized in dimethyl sulfoxide in this second species. Safety and pharmacokinetic studies were conducted in naïve dogs. After confirming safety, subsequent pharmacokinetic analyses demonstrated that a 100 mg/kg subcutaneous dose of GM6001 resulted in plasma concentrations that peaked shortly after administration and were sustained for at least 4 days at levels that produced robust in vitro inhibition of matrix metalloproteinase-9. A randomized, blinded, placebo-controlled study was then conducted to assess efficacy of GM6001 given within 48 hours of spinal cord injury. Dogs were enrolled in 3 groups: GM6001 dissolved in dimethyl sulfoxide (n = 35), dimethyl sulfoxide (n = 37), or saline (n = 41). Matrix metalloproteinase activity was increased in the serum of injured dogs and GM6001 reduced this serum protease activity compared to the other two groups. To assess recovery, dogs were a priori stratified into a severely injured group and a mild-to-moderate injured group, using a Modified Frankel Scale. The Texas Spinal Cord Injury Score was then used to assess long-term motor/sensory function. In dogs with severe spinal cord injuries, those treated with saline had a mean motor score of 2 (95% CI 0-4.0) that was significantly (P<0.05; generalized linear model) less than the estimated mean motor score for dogs receiving dimethyl sulfoxide (mean, 5; 95% CI 2.0-8.0) or GM6001 (mean, 5; 95% CI 2.0-8.0). As there was no independent effect of GM6001, we attribute improved neurological outcomes to dimethyl sulfoxide, a pleotropic agent that may target diverse secondary pathogenic events that emerge in the acutely injured cord.

PubMed Disclaimer

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Pharmacokinetics of GM6001 in dogs.
Administration of a single 100 mg/kg subcutaneous dose of GM6001 to dogs resulted in the rapid development of peak plasma drug concentrations with drug still detectable 96 hours post-delivery. Administration of a second dose of GM6001 to a sub-group of dogs 12 hours following initial drug delivery resulted in increased plasma drug concentrations at all assessed time points.
Figure 2
Figure 2. In vitro inhibition of MMP-9 by GM6001.
Calibrated MMP-9 activity (Log 106), as measured by a fluorescent electrophoretic technique, was dramatically attenuated by GM6001 in vitro at various concentrations. Plasma concentrations of GM6001, measured 96 hours following a single 100 mg/kg subcutaenous dose (range 0.16−.26 µM or 60–100 ng/mL), approximated those needed in vitro to robustly inhibit MMP-9 (0.2 µM or 77 ng/mL). Groups marked with an asterisk (*) had significantly (P<0.05) different calibrated MMP-9 activity from reference (no GM6001) using a one tailed Student’s t-test.
Figure 3
Figure 3. Consolidated Standards of Reporting Trials (CONSORT) Diagram.
Flow diagram depicting progress through different phases of the clinical trial including enrollment, group allocation, and follow-up.
Figure 4
Figure 4. T2-weighted magnetic resonance images in dogs with spinal cord injuries from intervertebral disk herniation.
In 1 dog (A, B) that was non-ambulatory with intact pelvic limb movement and sensation, there was focal ventrolateral spinal cord compression at the T12-T13 vertebral articulation without spinal cord signal change. A second dog (C, D) with paraplegia and absent pelvic limb deep nociception had compression at the T12-T13 vertebral articulation. There was extensive spinal cord T2-weighted hyperintensity (white arrows) visible on the sagittal image (C), suggestive of processes seen in contusion injuries such as edema, necrosis, hemorrhage, or cellular infiltrates. The transverse image (D, level of T13 vertebral body) indicated that T2-weighted hyperintensity was predominantly localized to the gray matter.
Figure 5
Figure 5. Serum MMP-2/MMP-9 activity in healthy and injured dogs.
Box-and-whisker plots summarizing the distribution of MMP 2/9 activity for healthy control dogs (N = 5) and dogs with spinal cord injury (SCI; N = 42) that had serum collected. Values of serum MMP 2/9 activities were significantly (P = 0.0128) greater for dogs with SCI included in the trial than control dogs. The horizontal lines with triangles represent the median value; the horizontal lines at the bottom and top of the boxes represent the 25th and 75th percentiles of the data, respectively. The thin vertical lines extending up or down from the boxes to horizontal lines (so-called whiskers) extend to a multiple of 1.75× the distance of the upper and lower quartile, respectively. Horizontal lines with circles represent values outside the limits of the whiskers.
Figure 6
Figure 6. Serum MMP-2/MMP-9 activity pre- and post-drug delivery.
Dogs were first randomized into 3 treatment groups, and serum was obtained prior to administration of saline, DMSO, or GM6001 (panel A). There were no differences in serum MMP-2/MMP-9 activity among treatment groups at the time of admission (panel A); however, at day 3, there was a significant (P = 0.0482) difference in serum activity between GM6001-treated dogs and the other treatment groups (panel B). See Figure 5 for a description of box-and-whisker plots. Groups marked with different letters differ significantly (P
Figure 7
Figure 7. Evaluation of primary outcome in dogs with SCI.
Box-and-whisker plots of TSCIS on day 42 by treatment group, stratified by MFS at admission (MFS = 0, left panel; or MFS >0, right panel). There were no significant differences in TSCIS among dogs with MFS score >0 (right panel), but TSCIS was significantly (P
Figure 8
Figure 8. Evaluation of TSCIS motor score at day 42 following SCI.
Box-and-whisker plots of TSCIS motor score on day 42 by treatment group, stratified by MFS at admission (MFS = 0, left panel; or MFS >0, right panel). There were no significant differences in motor score among dogs with MFS >0 (right panel), but motor score was significantly (P
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Zhang H, Adwanikar H, Werb Z, Noble-Haeusslein LJ (2010) Matrix metalloproteinases and neurotrauma: evolving roles in injury and reparative processes. Neuroscientist 16: 156–170. - PMC - PubMed
    1. Noble LJ, Donovan F, Igarashi T, Goussev S, Werb Z (2002) Matrix metalloproteinases limit functional recovery after spinal cord injury by modulation of early vascular events. J Neurosci 22: 7526–7535. - PMC - PubMed
    1. Wells JE, Rice TK, Nuttall RK, Edwards DR, Zekki H, et al. (2003) An adverse role for matrix metalloproteinase 12 after spinal cord injury in mice. J Neurosci 23: 10107–10115. - PMC - PubMed
    1. Zhang H, Trivedi A, Lee JU, Lohela M, Lee SM, et al. (2011) Matrix metalloproteinase-9 and stromal cell-derived factor-1 act synergistically to support migration of blood-borne monocytes into the injured spinal cord. J Neurosci 31: 15894–15903. - PMC - PubMed
    1. Shigemori Y, Katayama Y, Mori T, Maeda T, Kawamata T (2006) Matrix metalloproteinase-9 is associated with blood-brain barrier opening and brain edema formation after cortical contusion in rats. Acta Neurochir Suppl 96: 130–133. - PubMed

Publication types

MeSH terms

Substances