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Journal of Neurotrauma logoLink to Journal of Neurotrauma
. 2009 Jan;26(1):121–129. doi: 10.1089/neu.2008.0565

COG1410 Improves Cognitive Performance and Reduces Cortical Neuronal Loss in the Traumatically Injured Brain

Michael R Hoane 1,✉, Nicholas Kaufman 1, Michael P Vitek 2,,3,,4, Suzanne E McKenna 2
PMCID: PMC2749004  NIHMSID: NIHMS128106  PMID: 19119914

Abstract

We have previously shown that a single dose of COG1410, a small molecule ApoE-mimetic peptide derived from the apolipoprotein E (ApoE) receptor binding region, improves sensorimotor and motor outcome following cortical contusion injury (CCI). The present study evaluated a regimen of COG1410 following frontal CCI in order to examine its preclinical efficacy on cognitive recovery. Animals were prepared with a bilateral CCI of the frontal cortex. A regimen of COG1410 (0.8 mg/kg intravenously [IV]) was administered twice, at 30 min and again at 24 h post-CCI. Starting on day 11, the animals were tested for their acquisition of a reference memory task in the Morris water maze (MWM), followed by a working memory task in the MWM on day 15. Following CCI, the animals were also tested on the bilateral tactile adhesive removal test to measure sensorimotor dysfunction. On all of the behavioral tests the COG1410 group was no different from the uninjured sham group. Administration of the regimen of COG1410 significantly improved recovery on the reference and working memory tests, as well as on the sensorimotor test. Lesion analysis revealed that COG1410 significantly reduced the size of the injury cavity. Administration of COG1410 also reduced the number of degenerating neurons, as measured by Fluoro-Jade C staining, in the frontal cortex at 48 h post-CCI. These results suggest that a regimen of COG1410 appeared to block the development of significant behavioral deficits and reduced tissue loss. These combined findings suggest that COG1410 appears to have strong preclinical efficacy when administered following traumatic brain injury (TBI).

Key words: apoE, behavioral recovery, neuronal degeneration, neuroprotection, recovery of function, TBI, trauma

Introduction

Apolipoprotein E (ApoE: protein; APOE: gene) is the primary apolipoprotein synthesized in the brain in response to injury, where it functions via antioxidant, anti-inflammatory, anti-excitotoxic, and neurotrophic mechanisms (Nathan et al., 1994; Laskowitz et al., 1997a; Lomnitski et al., 1999; Laskowitz et al., 2001; Lynch et al., 2001; Colton et al., 2002; Aono et al., 2003). APOE has been shown to modulate the susceptibility and/or severity of neurological disorders characterized by neuroinflammation, including Alzheimer's disease, multiple sclerosis (MS), and CNS injury (Corder et al., 1993; Chapman et al., 2001; Zareparsi et al., 2002; Laskowitz and Vitek, 2007). However, the large ApoE protein has been shown to not cross the blood–brain barrier (Linton et al., 1991). The development of COG133, which corresponds to residues 133–149 from the entire ApoE protein, allows for brain penetration (Laskowitz et al., 2001). Previous work has demonstrated that this small-molecule peptide derived from amino acids 133–149 located in the receptor-binding region of ApoE (COG133) contains the neuroprotective, antioxidant, anti-excitotoxic, and anti-inflammatory properties of the intact ApoE holoprotein, in vitro and in vivo (Laskowitz et al., 2001; Misra et al., 2001; Aono et al., 2003; Lynch et al., 2003; Lynch et al., 2005; McAdoo et al., 2005). The in vivo neuroprotective activity of COG133 has been demonstrated in several clinically relevant models of neurological disorders, including TBI, MS, and hypoxic-ischemic injury (Lynch et al., 2005; McAdoo et al., 2005; Li et al., 2006). COG133 retains a measurable degree of alpha helicity present in the receptor-binding region of the native ApoE holoprotein from which it was derived (Laskowitz et al., 2001; Aono et al., 2003; Gay et al., 2006). Alanine scanning suggested that enhancing the alpha helical character of COG133 might enhance the anti-inflammatory activity, and this has subsequently led to the development of COG1410 (Laskowitz et al., 2006).

COG1410 is composed of apoE residues 138–149 with amino-iso-butyric acid substitutions at positions 140 and 145 to increase stability, and has exhibited superior anti-inflammatory efficacy and potency in vitro (Laskowitz et al., 2006). The in vivo efficacy was recently established in a model of subarachnoid hemorrhage (SAH) where treatment with COG1410 resulted in a dramatic reduction in mortality, vasospasm, cerebral edema, and functional deficits compared to vehicle-treated mice (Gao et al., 2006). The initial preclinical efficacy of a single dose of COG1410 was studied following unilateral CCI of the sensorimotor cortex. COG1410 at either low dose (0.4 mg/kg IV) or high dose (0.8 mg/kg IV) was administered 30 min post-CCI. It was found that the 0.8 mg/kg dose of COG1410 significantly improved sensorimotor performance and reduced lesion size (Hoane et al., 2007). A single-dose regimen of COG1410 administered 2 h following CCI in the mouse has also shown significant beneficial effects on behavioral outcome and neuroprotection (Laskowitz et al., 2007).

The purpose of the present study was to examine the preclinical efficacy of a two-dose regimen of COG1410 in the bilateral frontal CCI model. This model has been shown to be advantageous for the investigation of cognitive deficits following TBI (Lindner et al., 1998; Hoane et al., 2003), and has shown neuronal loss following injury in the thalamus and nucleus basalis magnocellularis (Smith et al., 2000). Multiple biochemical and cellular systems are required for proper cognitive functioning and any or all of them are damaged following brain trauma to the frontal lobes. COG1410 displays multiple activities including antioxidant, anti-inflammatory, anti-excitotoxic, and neurotrophic mechanisms (Nathan et al., 1994; Laskowitz et al., 1997b; Lynch et al., 2001; Colton et al., 2002; Aono et al., 2003; Lynch et al., 2003; Laskowitz and Vitek, 2007), all of which may contribute to improving cognitive recovery following TBI. These results support the idea that new and effective treatments for TBI should be directed at multiple targets/mechanisms related to the pathological mechanisms associated with TBI (Narayan et al., 2002). The purpose of the present study was to examine a regimen of COG1410 on cognitive/spatial ability in the MWM following trauma to the frontal cortex.

Methods

Subjects

Thirty-nine male Sprague-Dawley rats approximately 3 months old were used for this experiment. The behavioral study contained four groups of animals (CCI-COG1410, CCI-vehicle, sham-vehicle, and sham-COG1410). Additionally, the acute anatomical study included four animals in each of the following groups (CCI-COG1410, CCI-vehicle, and sham-vehicle), harvested 48 h post-CCI. All experimental procedures were reviewed and approved by the Institutional Animal Care and Use Committee, and the study was conducted in a facility certified by the American Association for the Accreditation of Laboratory Animal Care. The rats were maintained on a standard 12 h light/dark cycle with food and water available ad libitum.

Surgery

The surgical procedure was performed using aseptic procedures and conditions. The CCI model utilized in the present study was originally based on the methods of Lindner and colleagues (Lindner et al., 1998), and adapted based on previous studies (Hoane et al., 2003; Hoane et al., 2005). Rats were anesthetized with isoflurane (3–5%) in oxygen (0.8 L/min) and then prepared for surgery. When the rat was unresponsive (no ocular or pedal reflexes) the head was shaved and then scrubbed with 70% alcohol followed by povidone-iodine and placed into a digital stereotaxic device (Stoelting Instruments, Wood Dale, IL). A midline incision was made in the skin and underlying fascia. A circular craniotomy (6.0 mm) was performed using a surgical drill and a specially designed drill bit that prevented damage to the meninges and cortex. The craniotomy was bilateral and centered over the frontal cortex (3.0 mm anterior to the bregma). The contusion injury was created with an electromagnetic contusion device (http://myneurolab.com) using a sterile, stainless steel impactor tip (5.0 mm diameter) that was activated at a velocity of 2.78 m/sec. The impactor tip was positioned above the cortex and upon activation of the piston, the impactor tip made contact with the cortex for 0.5 sec, which resulted in a 2.0 mm compression of the cortex. Following the contusion any bleeding was controlled by brief application of sterile sponges soaked in cold 0.9% saline and the incision was closed with nylon suture material. The wound was treated with triple antibiotic ointment. Normal body temperature (37°C) was maintained with a warm water recycling pump and bed system (EZ-Anesthesia, Inc., Palmer, PA) during the surgical procedure and recovery. Rats receiving sham surgeries were treated in an identical manner to those receiving injuries; however, no CCI was performed.

Synthesis of peptides

Peptides were synthesized by NeoMPS (San Diego, CA) to a purity of 95%. COG1410 is Ac- AS(Aib)LRKL(Aib)KRLL-amide, which is derived from apoE residues 138–149 with Aib substitutions at positions 140 and 145. The peptides were reconstituted in a sterile, 0.9% saline solution at a dose of 0.8 mg/kg prior to administration (Hoane et al., 2007).

Drug administration

The rats were randomly assigned to either the COG1410 (0.8 mg/kg IV) or vehicle (0.9% saline IV) groups (Hoane et al., 2007). Injections were given 30 min and 24 h following injury via tail vein infusion. Infusions were performed under brief (<5 min), light isoflurane anesthesia (1–3%). All testing and analyses were conducted without knowledge of group assignment.

Cognitive assessment

The MWM has been widely utilized to assess cognitive performance following brain injury. A blue fiberglass tank, 1.5 m in diameter and 76 cm deep, was filled with water to a depth of 32 cm at 24°C. A 10-cm2 acrylic glass platform was submerged 1.0 cm below the surface of the water. Path length and latency to escape were recorded by a computerized video tracking system and SMART tracking system (San Diego Instruments, San Diego, CA). All the animals were assessed on the acquisition of a reference memory task beginning on days 11–14 post-CCI (Hoane et al., 2003; Hoane et al., 2006b). Each rat was given four trials a day, starting from one of four release points chosen at random. The trial was terminated when the rat reached the submerged platform located in the center of the northeast quadrant, or when 90 sec had elapsed. Each rat was allowed to remain on the platform for 30 sec, after which it was placed in a warm holding cage for 15 min before the next trial.

Working memory performance was tested on days 15–18 post-CCI using established methods (Lindner et al., 1998; Hoane et al., 2003; Hoane, 2005). The platform was submerged at the center of a new randomly chosen quadrant (southwest, northwest, and southeast) each day. Each animal was given four trials per day, starting from one of four randomly chosen release points (inter-trial interval [ITI] =15 min). The first trial on each of these three days was considered an information trial and was not included in subsequent analyses. Each trial ended when the animal located the platform, or when 90 sec had elapsed. Although, a 15 min ITI was used in the present working memory paradigm, other studies have used shorter and longer ITIs in rodents, some reaching several hours in length (Means, 1993; Means et al., 1996; Kline et al., 2002; Fitch et al., 2008; Schulteis et al., 2008).

Bilateral tactile adhesive removal test

Somatosensory dysfunction was assessed using this test, in which the latency time to remove a small round adhesive patch (113 mm2) from the radial aspect of each forelimb was recorded (Hoane et al., 2003; Hoane et al., 2005; Schallert and Woodlee, 2005). Two trials were administered 5 min apart to minimize habituation effects. Each trial was terminated when both patches had been removed or at the end of 2 min. Baseline latencies to remove the patches were recorded prior to injury, after which each rat was tested on days 2, 4, 6, 8, 10, and 14 post-CCI.

Histology

At 22 days post-CCI, the rats in the behavioral study were anesthetized with urethane (3.0 g/kg intraperitoneally) and transcardially perfused with 0.9% phosphate-buffered saline (PBS), followed by 10% phosphate-buffered formalin (PBF). The brains were post-fixed in PBF following removal from the cranium. A 30% sucrose solution was used to cryopreserve the brains for 3 days prior to frozen sectioning. Serial sections (40 μm thick) were sliced using a sliding microtome with an electronic freezing stage and collected and placed in PBS. The rats in the anatomical study were perfused 48 h following CCI and the brains were processed for paraffin embedding. The brains were processed with an automated tissue processor using a standard protocol for rodent brain (Holland et al., 2008). Paraffin-embedded brains were sectioned at 8 μm on a Leica RM2125 rotary microtome (Leica Microsystems, Bannockburn, IL), and serial sections were mounted on microscope slides. Immediately prior to staining, the slides were deparaffinized in two 7-min washes of xylene, then transferred through a series of EtOH baths (2 × 5 min in 100%; 3 min each at 95%, 70%, and 50%), followed by a 1-min rinse in distilled H2O (dH2O).

Lesion analysis

Following frozen slicing, a series of coronal sections, mounted on gelatin-subbed microscope slides, were stained with cresyl violet, dehydrated, and cover-slipped. The extent of the lesion was analyzed with an Olympus microscope (BX-51; Olympus, Center Valley, PA) and DP-70 camera. Images of the sections throughout the extent of the injury were captured using the digital capturing system, and measurement of area of the lesioned tissue was determined using University of Texas Health Science Center San Antonio (UTHSCSA) ImageTool software (Barbre and Hoane, 2006). The Calvalieri method was used to calculate the volumes of the remaining intact and healthy frontal cortex (Coggeshall, 1992). The number of sections and the section thickness (40 μm) were multiplied by the mean area of the lesion cavity (calculated at four stereotaxic coordinates surrounding the lesion: 3.00, 2.00, 1.00, and 0.00 relative to the bregma) (Paxinos and Watson, 2005). The extent of cortical injury was measured by calculating the volume of remaining cortex at the level of the injury (Hoane et al., 2003; Hoane et al., 2005).

Neuronal degeneration

Two series of deparaffinized sections at the middle of the injury cavity (1.5 and 2.5 mm anterior to the bregma) were stained for degenerating neurons using Fluoro-Jade C® (Chemicon Int., Billeria, MA) stain (FJ). Standard protocol was followed (Hoane et al., 2006a; Holland et al., 2008). The slides were then immersed in a 1% sodium hydroxide/80% EtOH solution for 5 min, and then transferred to 70% EtOH for 2 min, followed by a rinse in dH2O for 2 min. The slides were then transferred to a solution of 0.06% potassium permanganate for 10 min and agitated. The slides were then rinsed in dH2O for 2 min and transferred to the staining solution containing 0.004% FJ for 20 min in the dark. The slides were rinsed in dH2O (3 × 1 min) and then were allowed to air dry in the dark. The slides were then cleared in xylene for 1 min and cover-slipped. Tissue sections were visualized with an Olympus fluorescent microscope (BX-51) system using a GFP filter. A section was viewed at 4 × magnification, which was then increased to 40 × , and the image was captured using a 13-megapixel Olympus (DP-70) digital camera. The number of FJ+ neurons surrounding the lesion was determined using cell counts from three sites randomly selected from the tissue surrounding the injury site that were expressing fluorescence signal in each hemisphere. The number of FJ+ cells within the captured field of view (35.29 mm2) was counted using ImageTool software. The total number of FJ+ cells counted from the 12 different sites was used for the analyses.

Statistical analysis

Analysis of variance (ANOVA) tests were performed using procedures for general linear models (SPSS 15.0 for Windows; SPSS, Inc. Chicago, IL) with options for repeated measures, where appropriate, for all behavioral measures. Initial analyses were performed between the two sham groups (sham-vehicle [n = 7] and sham-COG1410 [n = 6]) on all of the behavioral tests and lesion analyses. These analyses revealed no significant differences between the sham groups, thus both groups were combined into one sham group for all subsequent analyses. The between-group factor was group (0.8 mg/kg COG1410 [n = 7], vehicle [n = 7], sham [n = 13]), and the within-group factor was day of testing. Huynh-Feldt probabilities and Fischer's least significant difference tests (LSD) were used to correct for type I errors associated with repeated measures and post-hoc means comparisons, respectively. To ensure correction for type I errors, all Huynh-Feldt probability corrections used the actual corrected degrees of freedom instead of the uncorrected degrees of freedom. Anatomical data were analyzed with one-way ANOVA procedures and Fischer's LSD. A significance level of p < 0.05 was used for all statistical analyses.

Results

Sham analysis

To determine the effect of administration of COG1410 on normal behavior on the reference memory test in the MWM, a one-way repeated measures ANOVA, with group (sham-vehicle and sham-COG1410) and day (11, 12, 13, and 14) as the factors in the analysis was performed. The analysis showed no significant difference in performance, and the main effect of group was not significant [F(1, 11) = 0.24, p > 0.64], nor was the day x group interaction [F(2.89, 31.86) = 0.28, p > 0.83]. However, there was a significant effect for days, suggesting that both groups improved over time [F(2.89, 31.86) = 37.22, p < 0.001].

The effect of administration of COG1410 on normal behavior on the working memory test in the MWM was analyzed with a one-way repeated measures ANOVA, with group (sham-vehicle and sham-COG1410) and day (15, 16, 17, and 18) as the factors in the analysis. The analysis showed no significant difference in performance, and the main effect of group was not significant [F(1, 11) = 0.99, p > 0.34], nor was the day x group interaction [F(1.46, 16.01) = 1.16, p > 0.32]. However, there was a significant effect for days, suggesting that both groups improved over time [F(1.46, 16.01) = 7.09, p < 0.01].

The effect of administration of COG1410 on normal behavior on the bilateral tactile adhesive removal test was analyzed with a one-way repeated measures ANOVA, with group (sham-vehicle, sham-COG1410) and day (0, 2, 4, 6, 8, 11, and 14) as the factors in the analysis. The analysis showed no significant difference in performance, and the main effect of group was not significant [F(1, 11) = 0.29, p > 0.60], nor was the day x group interaction [F(3.53, 38.82) = 0.66, p > 0.60]. There was also a non-significant effect for days [F(3.53, 38.82) = 0.50, p > 0.72].

There was also no difference in remaining cortical volume between the two sham groups. A one-way ANOVA, with group (sham-vehicle or sham-COG1410) was performed. The effect of group was not significant [F(1, 11) = 1.65, p > 0.23].

MWM: Acquisition of reference memory

Post-CCI performance on the acquisition of a reference memory task was assessed with a one-way repeated measures ANOVA, with group (0.8 mg/kg COG1410 and sham-vehicle) and day (11, 12, 13, and 14) as the factors in the analysis. The analysis showed a significant improvement in performance over time, and the main effect for days was significant [F(2.44, 58.62) = 63.13, p < 0.001]. There was a significant difference in behavioral performance between groups, and the main effect for group was significant [F(2, 24) = 7.06, p < 0.004] (Fig. 1). However, the recovery did not differ significantly across time, and the day x group interaction was not significant [F(4.52, 40.67) = 0.65, p > 0.65]. Post-hoc analysis of the significant main effect of group revealed that the 0.8-mg/kg COG1410 group showed improved performance compared to the vehicle group collapsed over testing days [LSD (12) = 13.78, p < 0.03] (Fig. 1, total latency graph). Post-hoc analysis also revealed that the 0.8-mg/kg COG1410 group was not significantly different across time compared to the sham group [LSD (18) = 5.56, p > 0.30].

FIG. 1.

FIG. 1.

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The effects of a regimen of COG1410 (0.8 mg/kg IV) or vehicle administered following frontal CCI or sham surgery on the acquisition of reference memory in the MWM. The left graph shows the plotted mean (±SEM) swim latencies to the submerged platform over days. The overall main effect of group (collapsed over days) is graphically shown on the right. Treatment with COG1410 significantly reduced the reference memory acquisition deficits compared to the vehicle group (* = p < 0.05), and was not significantly different from the sham group (^p > 0.05).

MWM: Working memory performance

Post-CCI performance on working memory was assessed with a one-way repeated measures ANOVA, with group (0.8 mg/kg COG1410 and sham-vehicle) and day (15, 16, 17, and 18) as the factors in the analysis. The analysis showed a significant improvement in performance over time, and the main effect for days was significant [F(3.00, 72.00) = 5.65, p < 0.002]. There was a significant difference in behavioral performance between groups, and the main effect for group was also significant [F(2, 24) = 4.65, p < 0.02] (Fig. 2). However, the recovery did not differ significantly across time, and the day x group interaction was not significant [F(6.00, 72.00) = 1.03, p > 0.41]. Post-hoc analysis of the significant main effect of group revealed that the 0.8-mg/kg COG1410 group showed improved performance compared to the vehicle group across testing days [LSD (12) = 12.80, p < 0.03] (Fig. 2, total latency graph). Post-hoc analysis also revealed that the 0.8-mg/kg COG1410 group was not significantly different across time compared to the sham group [LSD (18) = 1.51, p > 0.80].

FIG. 2.

FIG. 2.

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The effects of a regimen of COG1410 (0.8 mg/kg IV) or vehicle administered following frontal CCI or sham surgery on the working memory task in the MWM. The left graph shows the plotted mean (±SEM) swim latencies to the submerged platform over days. The overall main effect of group (collapsed over days) is graphically shown on the right. Treatment with COG1410 significantly reduced the working memory deficit compared to the vehicle group (* = p < 0.05) and was not significantly different from the sham group (^p > 0.05).

Bilateral tactile adhesive removal test

Post-CCI performance on the latencies to remove the tactile stimuli were assessed with a one-way repeated measures ANOVA, with group (0.8 mg/kg COG1410 and sham-vehicle) and day (0, 2, 4, 6, 8, 11, and 14) as the factors in the analysis. The analysis showed a significant improvement of performance over time, and the main effect for days was significant [F(3.30, 79.11) = 16.49, p < 0.001]. There was a significant difference in behavioral performance between groups, and the main effect for group was also significant [F(2, 24) = 5.96, p < 0.008]. The recovery differed significantly across time, and the day x group interaction was significant [F(6.59, 79.11) = 10.241, p < 0.001] (Fig. 3). Post-hoc analysis of the significant interaction revealed that the 0.8-mg/kg COG1410 group showed improved performance compared to the vehicle group on post-CCI days 2 [LSD (12) = 44.30, p < 0.002], 4 [LSD (12) = 29.71, p < 0.006], and 6 [LSD (12) = 15.07, p < 0.04]. Post-hoc analysis revealed that the 0.8-mg/kg COG1410 group was not significantly different compared to the sham group on any test day (p > 0.05).

FIG. 3.

FIG. 3.

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The effects of a regimen of COG1410 (0.8 mg/kg IV) or vehicle administered following frontal CCI or sham surgery on the bilateral tactile removal test. The graph shows the plotted mean (±SEM) latencies to remove the stimuli from both forelimbs. Treatment with COG1410 significantly reduced the magnitude of the injury compared to the vehicle group (* = p < 0.05) and was not significantly different from the sham group (^ = p > 0.05).

Lesion analysis

The reduction in cortical tissue volume through the injury site was analyzed with a one-way ANOVA with group (0.8 mg/kg COG1410 and sham-vehicle) as the factor in the analysis. There was a significant difference in injury volume between groups [F(2, 24) = 4.15, p < 0.03] (Fig. 4). Post-hoc analysis of the significant main effect of group revealed that the 0.8-mg/kg COG1410 group showed significantly less cortical tissue loss compared to the vehicle group [LSD (12) = 0.78, p < 0.04]. Post-hoc analysis also revealed that the loss of cortical tissue in the 0.8-mg/kg COG1410 group was not significantly different compared to the sham group [LSD (18) = 0.07, p > 0.82].

FIG. 4.

FIG. 4.

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Lesion analysis. Plotted is the mean (±SEM) remaining cortical volume for each group. COG1410 (0.8 mg/kg) reduced the amount of injury-induced tissue loss compared to the vehicle group (* = p < 0.05). The remaining tissue volume in the COG1410-treated group was not significantly different from the sham group (^ = p > 0.05).

Representative histological images are provided in Fig. 5.

FIG. 5.

FIG. 5.

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Histology plate. Shown are representative cresyl violet images of sections (3.20, 2.20, and 1.20 mm relative to the bregma) from a vehicle-treated brain (remaining cortical volume = 9.75 mm3, average group volume = 9.68 mm3), and a representative COG1410-treated brain (remaining cortical area = 10.80 mm3, average group volume = 10.46 mm3) (0.44 × magnification; scale bar = 2.0 mm).

Neuronal loss

The reduction in the number of dying neurons (FJ+) at the injury site was analyzed with a one-way ANOVA with group (0.8 mg/kg COG1410 or sham-vehicle) as the factor in the analysis. There was a significant difference in neuronal loss between groups [F(2,11) = 23.34, p < 0.001]. Post-hoc analysis revealed that the 0.8-mg/kg COG1410 group showed significantly less neuronal loss compared to the vehicle group [LSD (6) = 89.75, p < 0.002] (Fig. 6).

FIG. 6.

FIG. 6.

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Acute neuroprotection. Plotted is the mean (±SEM) number of FJ+ neurons in the injured cortex at 48 h post-CCI for each group. Administration of COG1410 (0.8 mg/kg) significantly reduced the number of FJ+ neurons compared to the vehicle group (* = p < 0.05).

Discussion

The results of this study have shown that a small-molecule peptide derived from ApoE has therapeutic benefit when administered following induction of TBI. COG1410 exhibited strong effects on recovery of function and neuroprotection following frontal CCI. Specifically, the 0.8-mg/kg dose of COG1410 induced recovery in the MWM cognitive tests. The regimen of COG1410 significantly facilitated the acquisition of the reference memory task in the MWM compared to the vehicle-treated group. Furthermore, the COG1410-treated group and the sham control group were statistically indistinguishable. The working memory task was conducted beginning on day 15 post-CCI, and also demonstrated that COG1410 significantly improved working memory performance compared to the vehicle-treated group. In fact, it appears that the administration of COG1410 also prevented the occurrence of the working memory deficit.

Beneficial effects were also seen on the bilateral tactile adhesive removal test. COG1410 significantly reduced the magnitude of the injury deficit beginning on post-CCI day 2, and reached a recovery plateau by day 4. There was a greater than 50% reduction in injury impairment in the COG1410 group on day 2 compared to the vehicle group. Interestingly, the treatment group also was not significantly different from the sham controls, suggesting that there was an abatement of the injury-induced deficit.

In a similar manner to what was seen with the behavioral data, the anatomical data from the present study also showed significant neuroprotective effects. The 0.8-mg/kg dose of COG1410 significantly reduced cortical cavity development following CCI. The analysis of the remaining frontal cortex volume showed that the vehicle-treated group had the lowest remaining volume (9.68 mm3) compared to that of the 0.8-mg/kg dose of COG1410 (10.46 mm3) and the sham cortex volume (10.53 mm3). Thus COG1410 significantly prevented cortical loss following frontal CCI. This effect is similar to what we have shown following a single treatment of COG1410 following unilateral CCI (Hoane et al., 2007). In the present study, we also conducted an acute neuroprotection study. Groups of animals were sacrificed 48 h after frontal CCI and the brains were processed for FJ reactivity to label degenerating neurons. It was found that COG1410 reduced the expression of FJ within the injured frontal cortex compared to vehicle control. The COG1410 group showed an average of 46 FJ+ neurons, compared to 136 FJ+ neurons in the vehicle group; this resulted in a reduction of 34%. A similar neurprotective effect has been seen with COG1410 in the hippocampus following TBI (Laskowitz et al., 2007).

The ability of ApoE to reduce glial activation and modulate CNS inflammation has been previously demonstrated (Laskowitz et al., 1997a; Laskowitz et al., 2001; Lynch et al., 2003). In a previous study, COG1410 was found to modulate reactive gliosis following TBI. A single injection of COG1410 following unilateral CCI significantly reduced the number and size of GFAP+ cells in the injured cortex (Hoane et al., 2007). Thus, ApoE-derived compounds (i.e., COG133 and COG1410) have glial and inflammatory modulating capabilities in vivo. It has also been previously shown that COG1410 reduced cerebral edema in a murine model of SAH (Gao et al., 2006). The neuroprotective effects of ApoE mimetic peptides in preclinical models of TBI, SAH, hypoxic-ischemic injury, and MS suggest that ApoE-mimetic peptides represent a novel therapy for the treatment of neurological diseases with neuroinflammatory components (Lynch et al., 2005; Gao et al., 2006; Hoane et al., 2007; Laskowitz et al., 2007; Laskowitz and Vitek, 2007).

The results of this study indicate that a two-dose regimen of COG1410 significantly improved performance on all behavioral measures of sensorimotor and cognitive function. This effect was very strong, actually appearing to eliminate most behavioral deficits. COG1410 also reduced tissue loss and provided significant neuroprotection in the injured cortex. The behavioral effects seen with this regimen of COG1410 (two administrations) appears much stronger than what we have previously shown following a single administration of COG1410 (Hoane et al., 2007). It is likely that a regimen that includes increasing the frequency of dosing will further increase the preclinical effectiveness. This study supports the continued preclinical examination of COG1410 as a treatment for TBI.

Acknowledgments

The research was supported by Cognosci Inc. and by grants R44NS048689 and R44AG020473 from the National Institutes of Health. Dr. Vitek is an Associate Professor of Neurology and an Adjunct Professor of Neurobiology at Duke University Medical Center, and is also Chief Executive Officer of Cognosci, Inc. Cognosci, Inc. has filed for patent protection for COG1410.

Author Disclosure Statement

No conflicting financial interests exist.

References

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