Benzoates for the treatment of nervous system injuries and disorders

Abstract

Salts of benzoic acid and prodrugs thereof for slowing the progression or reducing the severity of symptoms associated with nervous system injury in a subject are disclosed.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 63 / 290,633, filed December 16, 2021, which is incorporated by reference in its entirety. [Background technology]

[0002] Each year, approximately 1.7 million people in the United States and 10 million people worldwide suffer from traumatic brain injury (TBI). TBI is the leading cause of death and disability in the United States, contributing to approximately 30% of all injury-related deaths. While all people are at risk, military personnel are at higher risk for TBI due to the nature of their occupation. During TBI, chronic neuroinflammation and demyelination and / or impaired remyelination contribute significantly to disability among military personnel. Although treatments exist to reduce or eliminate certain physical, emotional, and cognitive problems associated with TBI, effective neuroprotective therapies are needed for people to recover from TBI.

[0003] Following TBI and other nervous system injuries, a complex series of pathophysiological events occur, causing both structural damage and functional deficits. Activation of glial cells in the nervous system and upregulation of associated pro-inflammatory molecules are involved in the pathogenesis of several neurodegenerative and neuroinflammatory diseases. Thus, one of the main features of both acute and chronic TBI is also neuroinflammation, which is evidenced within minutes of TBI. Studies from experimental animals with focal and diffuse TBI have shown that various pro-inflammatory molecules, such as IL-1β, TNF-α, and inducible nitric oxide synthase (iNOS), are involved in the pathogenesis of TBI. Many clinical studies have shown increased IL-1β and TNF-α in the CSF and serum of TBI patients compared to healthy controls. Upregulation of a broad spectrum of pro-inflammatory molecules in the brain causes edema, blood-brain barrier (BBB) ​​leakage, neuronal apoptosis, and atrophy, ultimately leading to functional impairment. There is a need for more effective treatments for nervous system injuries, including TBI. Summary of the Invention

[0004] In one aspect, a method of slowing the progression or reducing the severity of symptoms associated with nervous system injury in a subject in need thereof comprises administering to the subject an effective amount of a benzoate salt or a prodrug thereof, thereby slowing the progression or reducing the severity of symptoms associated with nervous system injury.

[0005] In another aspect, a method of slowing the progression or reducing the severity of symptoms associated with nervous system injury in a subject in need thereof comprises administering to the subject an effective amount of sodium benzoate, thereby slowing the progression or reducing the severity of symptoms associated with nervous system injury.

[0006] The foregoing summary, and the following description of the present disclosure, can be better understood when read in conjunction with the accompanying drawings. For the purpose of illustrating the present disclosure, the drawings illustrate some, but not all, alternative embodiments. The disclosure is not limited to the precise arrangements and instrumentalities shown. The following figures, which are incorporated in and constitute a part of this specification, serve to explain the principles of the present disclosure. [Brief description of the drawings]

[0007] [Figure 1]FIG. 1A is a photograph of the tip of the CCI device with the mouse brain exposed. Using the CCI technique, brain injury was gently induced in the exposed brain region of anesthetized mice. FIG. 1B is a photograph showing the induced CCI in the brain. Blood clots and tissue damage at the burr hole (stereotaxic coordinates - 1.5 mm posterior and 1.5 mm lateral from bregma) were found in the injured brain region of the mouse after CCI injury. FIG. 1C is an image showing the induction of mild, moderate, and severe CCI injury using a 1 mm tip at three different speeds, namely 1.0 V, 1.25 V, and 1.5 V, respectively. One week after injury, mice (n=3) were perfused with 4% paraformaldehyde, followed by removal of the brain and staining of brain sections with cresyl violet. FIG. 1D is a schematic diagram of the experimental design showing the course of treatment, behavior, and histological analysis after CCI injury (1 mm tip / 1.0 V). [Diagram 2]FIG. 2A is an image of double-labeled immunofluorescence for GFAP and iNOS in brain sections for control. Mice were treated with 50 mg / kg / day NaB or NaFO by oral administration after induction of CCI injury. After 7 days of NaB treatment, the brain sections were analyzed by double-labeled immunofluorescence for GFAP and iNOS. These results show that oral treatment of NaB attenuates the activation of astrocytes in vivo in the cortical and hippocampal regions of mice with CCI injury. FIG. 2B is an image of double-labeled immunofluorescence for GFAP and iNOS in brain sections for CCI injury. Mice were treated with 50 mg / kg / day NaB or NaFO by oral administration after induction of CCI injury. After 7 days of NaB treatment, the brain sections were analyzed by double-labeled immunofluorescence for GFAP and iNOS. These results show that oral treatment of NaB attenuates the activation of astrocytes in vivo in the cortical and hippocampal regions of mice with CCI injury. FIG. 2C is an image of double-labeled immunofluorescence for GFAP and iNOS in brain sections for CCI+NaB. After induction of CCI injury, mice were treated with 50 mg / kg / day NaB or NaFO by oral administration. After 7 days of NaB treatment, brain sections were analyzed by double-labeled immunofluorescence for GFAP and iNOS. These results show that oral treatment of NaB attenuates in vivo activation of astrocytes in the cortical and hippocampal regions of mice with CCI injury. FIG. 2D is an image of double-labeled immunofluorescence for GFAP and iNOS in brain sections for CCI+NaFO. After induction of CCI injury, mice were treated with 50 mg / kg / day NaB or NaFO by oral administration. After 7 days of NaB treatment, brain sections were analyzed by double-labeled immunofluorescence for GFAP and iNOS. These results show that oral treatment of NaB attenuates in vivo activation of astrocytes in the cortical and hippocampal regions of mice with CCI injury. Figure 2E is a histogram of cells positive for GFAP counted in the cortical region. Results represent an analysis of six sections of each of six mice per group. ap<0.001 vs. control, bp<0.001 vs. CCI injury. Figure 2F is a histogram of cells positive for GFAP counted in the CA1 region.Results represent an analysis of six sections of each of six mice per group. ap<0.001 vs. control, bp<0.001 vs. CCI injury. FIG. 2G is a histogram of cells positive for iNOS counted in the cortical region. Results represent an analysis of six sections of each of six mice per group. ap<0.001 vs. control, bp<0.001 vs. CCI injury. FIG. 2H is a histogram of cells positive for iNOS counted in the CA1 region. Results represent an analysis of six sections of each of six mice per group. ap<0.001 vs. control, bp<0.001 vs. CCI injury. FIG. 2I is an immunoblot image of tissue extracts of the hippocampal region from all mouse groups (n=4 per group) for GFAP. Actin was run as a loading control. FIG. 2J is a plot showing the values ​​of GFAP / actin relative to the control obtained by immunoblot band scanning. ap<0.001 vs. control, bp<0.001 vs. CCI injury. [Diagram 3]FIG. 3A is an image of double-labeled fluorescence for Iba1 and iNOS for control. Mice were treated with 50 mg / kg / day NaB or NaFO starting 24 hours after induction of CCI injury. After 7 days of treatment, brain sections were analyzed by double-labeled fluorescence for Iba1 and iNOS. These results show that NaB treatment inhibits in vivo microglial activation in the cortex and hippocampus of mice with CCI injury. FIG. 3B is an image of double-labeled fluorescence for Iba1 and iNOS for CCI injury. Mice were treated with 50 mg / kg / day NaB or NaFO starting 24 hours after induction of CCI injury. After 7 days of treatment, brain sections were analyzed by double-labeled fluorescence for Iba1 and iNOS. These results show that NaB treatment inhibits in vivo microglial activation in the cortex and hippocampus of mice with CCI injury. FIG. 3C is an image of double-labeled fluorescence for Iba1 and iNOS for CCI+NaB. Mice were treated with 50 mg / kg / day NaB or NaFO starting 24 hours after induction of CCI injury. After 7 days of treatment, brain sections were analyzed by double-label fluorescence for Iba1 and iNOS. These results show that NaB treatment inhibits in vivo microglial activation in the cortex and hippocampus of mice with CCI injury. Figure 3D is an image of double-label fluorescence for Iba1 and iNOS for CCI+NaFO. Mice were treated with 50 mg / kg / day NaB or NaFO starting 24 hours after induction of CCI injury. After 7 days of treatment, brain sections were analyzed by double-label fluorescence for Iba1 and iNOS. These results show that NaB treatment inhibits in vivo microglial activation in the cortex and hippocampus of mice with CCI injury. Figure 3E is a histogram of cells positive for Iba1 counted in the cortical region. Results represent the analysis of six sections of each of six mice per group. ap<0.001 vs. control, bp<0.001 vs. CCI injury. FIG. 3F is a histogram of cells positive for Iba1 counted in the CA1 region. Results represent an analysis of 6 sections each of 6 mice per group. ap<0.001 vs. control, bp<0.001 vs. CCI injury.FIG. 3G is an immunoblot image of tissue extracts of the hippocampal region from all mouse groups (n=4 per group) for Iba1. Actin was run as a loading control. FIG. 3H is a plot showing the values ​​of Iba1 / actin relative to the control obtained by immunoblot band scanning. ap<0.001 vs. control, bp<0.001 vs. CCI injury. FIG. 3I is an immunoblot image of tissue extracts of the hippocampal region from all mouse groups (n=4 per group) for iNOS. Actin was run as a loading control. FIG. 3J is a plot showing the values ​​of iNOS / actin relative to the control obtained by immunoblot band scanning. ap<0.001 vs. control, bp<0.001 vs. CCI injury. [Figure 4] Images of mouse corpus collosum showing levels of proteolipid protein (PLP) and A2B5, a marker for oligodendrocyte precursor cells (OPCs), in experimental conditions after 21 days. Oral administration of sodium benzoate (NaB), but not sodium formate (NaFO), stimulates remyelination in mice with traumatic brain injury (TBI). Mice (n=6 per group) had moderate TBI induced by controlled cortical impact (CCI). Two hours after TBI, mice were treated with oral gavage of NaB and NaFO (50 mg / kg body weight / day mixed with water). After 21 days of treatment, brain sections were double-labeled for PLP and A2B5. Results represent the analysis of two sections each of six mice per group. [Diagram 5]FIG. 5A is an image showing representative cresyl violet sections of mouse brains placed in a series of hippocampal regions showing the volume of the lesion cavity in various groups. NaB treatment reduces the lesion volume in mice with CCI injury. FIG. 5B is an exemplary image of a cresyl violet section. Note that the extent of damage induced in the brain was found to be reduced in mice treated with NaB when compared to untreated CCI mice and CCI injury mice treated with NaFo. FIG. 5C is a plot showing the lesion size in the experimental conditions. 21 days after injury, lesion size was quantitatively measured in control mice, untreated CCI injury mice, CCI mice treated with NaB, and CCI mice treated with NaFO. Statistical analysis was performed using Student's t-test on the injured side of the brain [ap<0.001 (5.623×10−7) vs. control, bp<0.001 (=0.001) vs. CCI injury]. [Figure 6]Figure 6A shows heat map analysis of mice with TBI induced by CCI (n=6 per group) in open field activity after 7 days of treatment. Open field activity was monitored by Ethovision XT13.0 open field activity system (Noldus). Two hours after TBI, mice were treated with oral gavage of NaB and NaFO (50 mg / kg body weight / day, mixed with water). Oral administration of sodium benzoate (NaB), but not sodium formate (NaFO), improves open field activity in mice with TBI. Figure 6B shows bar graphs showing distance traveled by mice with TBI induced by CCI (n=6 per group) in open field activity after 7 days of treatment. Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], speed [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6C is a bar graph showing the speed of mice with TBI induced by CCI (n=6 per group) in open field activity after 7 days of treatment. Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], speed [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6D is a bar graph showing center frequency of mice with TBI induced by CCI (n=6 per group) in open field activity after 7 days of treatment.Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], velocity [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], central frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6E is a bar graph showing the rearing behavior of mice with TBI induced by CCI (n=6 per group) in open field activity after 7 days of treatment. Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], velocity [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6F shows a heatmap analysis of mice with TBI induced by CCI (n=6 per group) in open field activity after 21 days of treatment. Open field activity was monitored by an Ethovision XT13.0 open field activity system (Noldus). Two hours after TBI, mice were treated with oral gavage of NaB and NaFO (50 mg / kg body weight / day, mixed with water). Figure 6G is a bar graph showing the distance traveled by mice with CCI-induced TBI (n=6 per group) in open field activity after 21 days of treatment.Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], speed [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6H is a bar graph showing the speed of mice with TBI induced by CCI (n=6 per group) in open field activity after 21 days of treatment. Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], speed [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6I is a bar graph showing center frequency of mice with TBI induced by CCI (n=6 per group) in open field activity after 21 days of treatment. Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI injury], speed [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI injury], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI injury] and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI injury]. Figure 6J is a bar graph showing rearing of mice with TBI induced by CCI (n=6 per group) in open field activity after 21 days of treatment.Statistical analysis was performed using Student's t-test for distance traveled [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0029) vs. CCI lesion], speed [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0078) vs. CCI lesion], center frequency [ap<0.001 (=0.0001) vs. control, bp<0.001 (=0.0036) vs. CCI lesion], and rearing behavior [ap<0.001 (=9.498x10-6) vs. control, bp<0.001 (=0.0081) vs. CCI lesion]. Figure 6K is a bar graph showing the results of the tail suspension test. After NaB treatment, mice with CCI injury showed significant improvement in the tail suspension test 7 days after injury [ap<0.001 (=5.725x10-8) vs. control, bp<0.001 (=0.0001) vs. CCI injury] and 21 days after injury [ap<0.001 (=3.995x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI injury]. Figure 6L is a bar graph showing the results of the rotarod test. The performance of mice with NaB treatment was significantly improved in the rotarod test 7 days after injury [ap<0.001 (=9.5998x10-9) vs. control, bp<0.001 (=0.0003) vs. CCI injury] and 21 days after injury [ap<0.001 (=1.2133x10-9) vs. control, bp<0.001 (=0.0010) vs. CCI injury]. Figure 6M is a bar graph showing the number of steps on the beam runway. Oral treatment with NaB also improved the performance of CCI-injured mice on the beam runway.Statistical significance was seen 7 days after injury [steps: ap<0.001 (=2.258x10-6) vs. control, bp<0.001 (=0.0027) vs. CCI injury, time taken: ap<0.001 (=2.979x10-7) vs. control, bp<0.001 (=0.0039) vs. CCI injury, leg slips: ap<0.001 (=1.567x10-7) vs. control, bp<0.001 (=0.0001) vs. CC injury] I-injury] and 21 days after injury [stepping: ap<0.001 (=0.0051) vs. control, ns (=0.2533) vs. CCI-injury, time taken: ap<0.001 (=0.0003) vs. control, ns (=0.1228) vs. CCI-injury, leg slip: ap<0.001 (=0.0001) vs. control, bp<0.05 (=0.0736) vs. CCI-injury] were performed using Student's t-test. Figure 6N is a bar graph showing time taken on the beam runway. Oral treatment with NaB also improved the performance of CCI-injured mice on the beam runway. Statistical significance was seen 7 days after injury [steps: ap<0.001 (=2.258x10-6) vs. control, bp<0.001 (=0.0027) vs. CCI injury, time taken: ap<0.001 (=2.979x10-7) vs. control, bp<0.001 (=0.0039) vs. CCI injury, leg slips: ap<0.001 (=1.567x10-7) vs. control, bp<0.001 (=0.0001) vs. CC injury] I-injury] and 21 days after injury [steps: ap<0.001 (=0.0051) vs. control, ns (=0.2533) vs. CCI-injury, time taken: ap<0.001 (=0.0003) vs. control, ns (=0.1228) vs. CCI-injury, leg slips: ap<0.001 (=0.0001) vs. control, bp<0.05 (=0.0736) vs. CCI-injury] were performed using Student's t-test. Figure 6O is a bar graph showing leg slips on the beam runway. Oral treatment with NaB also improved the performance of CCI-injured mice on the beam runway.Statistical significance was seen 7 days after injury [steps: ap<0.001 (=2.258x10-6) vs. control, bp<0.001 (=0.0027) vs. CCI injury, time taken: ap<0.001 (=2.979x10-7) vs. control, bp<0.001 (=0.0039) vs. CCI injury, leg slips: ap<0.001 (=1.567x10-7) vs. control, bp<0.001 (=0.0001) vs. CC injury] I-injury] and 21 days after injury [steps: ap<0.001 (=0.0051) vs. control, ns (=0.2533) vs. CCI-injury, time taken: ap<0.001 (=0.0003) vs. control, ns (=0.1228) vs. CCI-injury, leg slips: ap<0.001 (=0.0001) vs. control, bp<0.05 (=0.0736) vs. CCI-injury] were performed using Student's t-test. Figure 6P is a bar graph showing the number of steps on the grid runway. CCI-injured mice receiving NaB treatment showed improvement on the grid runway. Using Student's t-test, statistical significance was determined 7 days after injury [steps: ap<0.001 (=9.364x10-9) vs control, bp<0.001 (=3.394x10-5) vs CCI injury, time taken: ap<0.001 (=1.3770x10-7) vs control, bp<0.001 (=3.3886x10-5) vs CCI, leg placement: ap<0.001 (=2.737x10-8) vs. control, bp<0.001 (=5.954x10-6) vs. CCI injury] and 21 days after injury (steps: ap<0.001 (=0.0014) vs. control, bp<0.05 (=0.0718) vs. CCI injury, time taken: ap<0.001 (=5.562x10-5) vs. control, bp<0.05 (=0.072) vs. CCI injury, leg placement: ap<0.001 (=1.079x10-5) vs. control, ns (=0.2465) vs. CCI injury. ns-not significant. Figure 6Q is a bar graph showing time taken on the grid runway. CCI-injured mice receiving NaB treatment showed improvement on the grid runway.Using Student's t-test, statistical significance was determined 7 days after injury [steps: ap<0.001 (=9.364x10-9) vs. control, bp<0.001 (=3.394x10-5) vs. CCI injury, time taken: ap<0.001 (=1.3770x10-7) vs. control, bp<0.001 (=3.3886x10-5) vs. CCI, leg misplacement: ap<0.001 (=2.737x10-8) vs. control, bp<0.001 (=5. 954x10-6) vs. CCI injury] and 21 days after injury (steps: ap<0.001 (=0.0014) vs. control, bp<0.05 (=0.0718) vs. CCI injury; time taken: ap<0.001 (=5.562x10-5) vs. control, bp<0.05 (=0.072) vs. CCI injury; leg misplacement: ap<0.001 (=1.079x10-5) vs. control, ns (=0.2465) vs. CCI injury; ns-not significant). FIG. 6R is a bar graph showing leg placement on the grid runway. CCI-injured mice receiving NaB treatment showed improvement on the grid runway. Using Student's t-test, statistical significance was determined 7 days after injury [steps: ap<0.001 (=9.364x10-9) vs. control, bp<0.001 (=3.394x10-5) vs. CCI injury, time taken: ap<0.001 (=1.3770x10-7) vs. control, bp<0.001 (=3.3886x10-5) vs. CCI, leg placement: ap <0.001 (=2.737x10-8) vs. control, bp <0.001 (=5.954x10-6) vs. CCI injury] and 21 days after injury (steps: ap<0.001 (=0.0014) vs. control, bp <0.05 (=0.0718) vs. CCI injury; time taken: ap<0.001 (=5.562x10-5) vs. control, bp <0.05 (=0.072) vs. CCI injury; leg placement: ap<0.001 (=1.079x10-5) vs. control, ns (=0.2465) vs. CCI injury; ns-not significant). [Figure 7]Figure 7A shows heat maps depicting novel object recognition in mice with TBI induced by CCI (n=6 per group) 21 days after surgery. Starting 2 hours after TBI, mice were treated with NaB and NaFO (50 mg / kg body weight / day mixed with water) by oral gavage. Statistical analysis included novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI injury], Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI injury] and number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI injury], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI injury and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI injury] were performed by Student's t-test. ns-not significant. Figure 7B shows a heatmap showing the results of the Barnes circular maze test in mice with TBI induced by CCI (n=6 per group) 21 days after surgery. Statistical analysis included the novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI lesion], the Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI lesion], and the number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI injury], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI injury and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI injury] were performed by Student's t-test. ns-not significant. Figure 7C is a bar graph showing the exploration time of mice with TBI induced by CCI 21 days after surgery (n=6 per group) during the novel object recognition test.Statistical analysis included the novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI lesion], the Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI lesion], and the number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI injury], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI injury and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI injury] were performed by Student's t-test. ns-not significant. Figure 7D is a bar graph showing the latency of mice with TBI induced by CCI 21 days after surgery (n=6 per group) during the Barnes maze test. Statistical analysis included the novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI lesion], the Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI lesion], and the number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI injury], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI injury and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI injury] were performed by Student's t-test. ns-not significant. Figure 7E is a bar graph showing the number of errors of mice with TBI induced by CCI 21 days after surgery (n=6 per group) during the Barnes maze test.Statistical analysis included the novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI lesion], the Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI lesion], and the number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI injury], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI injury and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI injury] were performed by Student's t-test. ns-not significant. Figure 7F is a bar graph showing the number of positive turns of mice with TBI induced by CCI (n=6 per group) 21 days after surgery in the T-maze. Statistical analysis included the novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI lesion], the Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI lesion], and the number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI injury], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI injury and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI injury] were performed by Student's t-test. ns-not significant. Figure 7G is a bar graph showing the number of negative turns of mice with TBI induced by CCI (n=6 per group) 21 days after surgery in the T-maze.Statistical analysis included the novel object recognition test [exploration time: ap<0.001 (=2.5989x10-5) vs. control, bp<0.001 (=0.0003) vs. CCI lesion], the Barnes maze test [time taken: ap<0.001 (=1.7509x10-5) vs. control, bp<0.001 (=4.8824x10-5) vs. CCI lesion], and the number of errors: ap<0.001 (=3.234x10-5) vs. control]. , bp<0.001 (=0.0001) vs. CCI lesion], T-maze [positive turns: ap<0.001 (=3.3524x10-5) vs. control, bp<0.001 (=0.0004) vs. CCI lesion and negative turns: ap<0.001 (=3.3924x10-5) vs. control, bp<0.001 (=0.0005) vs. CCI lesion] performed by Student's t-test. ns - not significant. [Figure 8] Images of double-label immunofluorescence for GFAP and iNOS in control (FIG. 8A), CCI (FIG. 8B), CCI+GTB (FIG. 8C), and CCI+vehicle (FIG. 8D) brain sections. Mice were treated with 50 mg / kg / day GTB by oral gavage after induction of CCI injury. After 7 days of GTB treatment, brain sections were analyzed by double-label immunofluorescence for GFAP and iNOS. Cells positive for GFAP were counted in the cortex (FIG. 8E) and CA1 region (FIG. 8F) of the hippocampus. Similarly, cells positive for iNOS were also counted in the cortex (FIG. 8G) and CA1 region (FIG. 8H). Results represent the analysis of six sections each of six mice per group. Tissue extracts of the hippocampal region from all mouse groups (n=4 per group) were immunoblotted for GFAP (FIG. 8I) and iNOS (FIG. 8K). Actin was run as a loading control. Bands were scanned and values ​​(GFAP / actin) (FIG. 8J) and (iNOS / actin) (FIG. 8L) are presented relative to controls. These results show that oral administration of GTB inhibits in vivo astroglial inflammation in the cortex and hippocampus of mice with TBI. [Figure 9]Oral GTB reduces in vivo microglial activation in the cortex and hippocampus of mice with TBI. TBI was induced in mice by CCI injury, and 24 hours after injury, mice were treated with 50 mg / kg / day GTB by oral gavage. Seven days after GTB treatment, brain sections were double-labeled for Iba1 and iNOS (Figure 9A, control; Figure 9B, CCI; Figure 9C, CCI+GTB; Figure 9D, CCI+vehicle). Cells positive for Iba1 were counted in the cortex (Figure 9E) and CA1 regions (Figure 9F) of the hippocampus. Results represent the analysis of two sections each of six mice per group. Tissue extracts of the hippocampal region from all mouse groups (n=4 per group) were immunoblotted for Iba1 (Figure 9G). Actin was run as a loading control. Bands were scanned and values ​​(Iba1 / actin) (Figure 9H) are presented relative to the control. [Figure 10] Reduction of lesion volume in TBI mice by GTB treatment. TBI was induced in mice by CCI injury, and 24 hours after injury, mice were treated with 50 mg / kg / day GTB by oral gavage. Figure 10A. 21 days after injury, brain sections were stained with H&E, and the H&E stained sections were arranged in series to show the volume of the lesion cavity in different groups. Figure 10B shows an exemplary image of the H&E stained sections. Figure 10C. Lesion volume was quantified in all mouse groups. Statistical analysis was performed using two-way ANOVA, and lesion volumes were compared between the non-lesioned and lesioned sides of the brain, expressed as mean ± SD. [Figure 11]Restoration of PSD-95, NR2A, and GluR1 in the hippocampus of TBI mice by oral administration of GTB. TBI was induced in mice by CCI injury, and 24 hours after injury, mice were treated with 50 mg / kg / day GTB by oral gavage. 21 days after CCI injury, brain sections were double-labeled for NeuN and PSD-95 (Figure 11A, control; Figure 11B, CCI; Figure 11C, CCI+GTB; Figure 11D, CCI+vehicle). Results represent the analysis of one section each of six mice per group. Hippocampal tissue extracts from all mouse groups (n=4 per group) were immunoblotted for PSD-95, NR2A, and GluR1. Figure 11E. Actin was run as a loading control. Bands were scanned and values ​​(Iba1 / actin, FIG. 11F; NR2A / actin, FIG. 11G; GluR1 / actin, FIG. 11H) were presented relative to controls. Data are presented as mean + SD. Statistical analysis was performed using one-way ANOVA. [Figure 12] Effects of GTB on spatial learning and memory in TBI mice. TBI was induced in mice by CCI injury, and 24 hours after injury, mice were treated with 50 mg / kg / day GTB by oral gavage. 21 days after CCI injury, mice were tested by novel object recognition test (Figure 12A, heat map; Figure 12C, exploration time), Barnes maze (Figure 12B, heat map; Figure 12D, number of errors; Figure 12E, latency pr time), and T-maze (Figure 12F, positive turn; Figure 12G, negative turn). Six mice were used in each group. Statistical analysis was performed by one-way ANOVA followed by Tukey's post-hoc test. [Figure 13]GTB treatment restores motor function in TBI mice. TBI was induced in mice by CCI injury, and 24 hours after injury, mice were treated with 50 mg / kg / day GTB by oral gavage. Seven days after CCI injury, mice were tested for open field behavior (Figure 13A, heat map analysis monitored by using Noldus system; Figure 13B, distance traveled; Figure 13C, speed; Figure 13D, center frequency; Figure 13E, rearing), roto rod (Figure 13F, latency), tail suspension test (Figure 13G, immobility time), beam walking (Figure 13H, steps; Figure 13I, time taken; Figure 13J, slipping), and grid runway (Figure 13K, steps; Figure 13L, time taken; Figure 13M, misplacement). Six mice were used in each group. Statistical analysis was performed by one-way ANOVA followed by Tukey's post-hoc test. [Figure 14] Effects of GTB on motor function in TBI mice on day 21 after CCI injury. TBI was induced in mice by CCI injury, and 24 hours after injury, mice were treated with 50 mg / kg / day GTB by oral gavage. 21 days after CCI injury, mice were tested for open field behavior (Figure 14A, heat map analysis monitored by using Noldus system; Figure 14B, distance traveled; Figure 14C, speed; Figure 14D, center frequency; Figure 14E, rearing), rotorod (Figure 14F, latency), tail suspension test (Figure 14G, immobility time), beam walking (Figure 14H, steps; Figure 14I, time taken; Figure 14J, slipping), and grid runway (Figure 14K, steps; Figure 14L, time taken; Figure 14M, misplacement). Six mice were used in each group. Statistical analysis was performed by one-way ANOVA followed by Tukey's post-hoc test. [Figure 15]Effect of sodium benzoate (NaB) on the maturation of oligodendrocyte precursor cells (OPCs) into oligodendrocytes. OPCs were isolated from P1-P2 newborn mouse pups and cultured in OPC medium for 4 days in vitro (DIV), followed by treatment with 100 μM NaB and sodium formate (NaFO) in the absence of FGF and PDGF. Figure 15A) After 18 h of 100 μM NaB treatment in serum-free conditions, cells were fixed and then double immunostained with MBP (red) and OPC marker NG2 (green). Nuclei were stained with DAPI (blue). B) Similarly, OPCs were stained with PLP (red) and A2B5 (green) under similar treatment conditions. Quantification of MBP+ (Figure 15C), NG2+ (Figure 15D), PLP+ (Figure 15E), and A2B5+ (Figure 15F) cells as a percentage of total cells (DAPI+). Average of 5 fields per slide, 3 slides in total. Results are mean + SEM. ***p<0.001. G) For protein expression, cells were treated with various doses of NaB under serum-free conditions for 18 hours and then immunoblotted with PLP and MOG. Figure 15H) Densitometric analysis of bands for beta-actin was performed for PLP and MOG. ap<0.001 vs. control PLP, bp<0.005 vs. control MOG. Figure 15I) Cells were treated with NaB (100uM) and NaFO (100uM) under serum-free conditions for 4 hours and subsequently mRNA expression of myelin-specific genes was monitored by real-time PCR (ap<0.01 vs. control PLP, bp<0.01 vs. control MOG, cp<0.01 vs. control MBP, dp<0.01 vs. control CNPase). OPCs were cultured on top of randomly oriented polycaprolactone nanofibers (Nanofiber Solutions, Cat. No. Z694576) for 7 days. These cells were then treated with 100 μM NaB (FIG. 15J) and NaFO (FIG. 15K) for an additional 2 days, followed by immunofluorescence analysis for MBP (red) (FIG. 15J, control; FIG. 15K, NaB; FIG. 15L, NaFO). Images are displayed in a single red channel and merged with a phase contrast image of the nanofibers. A representative 3D reconstruction of OPCs attached to the nanofibers is also shown on the right. [Figure 16]Oral NaB stimulates in vivo maturation of OPCs in the corpus callosum of cuprizone-intoxicated mice. C57 / BL6 mice (8-10 weeks old, male) were fed a cuprizone-containing diet (Envigo) for 5 weeks and subsequently treated with NaB (50 mg / kg body weight / day) by oral gavage. (Figure 16A) After 3 weeks of treatment with NaB, corpus callosum sections were double-labeled for PLP and A2B5. Mean fluorescence intensity (MFI) of A2B5 (Figure 16B) and PLP (Figure 16C) was quantified from one section (two images per section) from each of the groups of five mice. Results are the mean + SEM of five mice per group. ***p<0.001, **p<0.01. [Figure 17] Effect of NaB on in vivo myelination in the corpus callosum of cuprizone-intoxicated mice. C57 / BL6 mice (8-10 weeks old, male, n=5) were fed a cuprizone-containing diet (Envigo) for 5 weeks and subsequently treated with NaB (50 mg / kg body weight / day) by oral gavage. After 3 weeks of treatment with NaB, corpus callosum sections were immunostained for MBP (Figure 17A) and PLP (Figure 17B). Mean fluorescence intensity (MFI) of MBP (Figure 17C) and PLP (Figure 17D) was quantified from one section (two images per section) from each of the groups of five mice. Results are the mean + SEM of five mice per group. **p<0.01, *p<0.05. Figure 17E) Corpus callosum sections were stained with luxol fast blue (LFB). FIG 17F) For electron microscopy studies, 50 μm thick sagittal sections were prepared and stained, followed by analysis of the corpus callosum sections for various parameters to evaluate the axonal ultrastructure. FIG 17G) G-score was calculated for all three groups, with 75 axons per group. FIG 17H) The percentage of myelinated axons was calculated in 7 randomly selected corpus callosum sections of 5 mice per group. ***p<0.001. [Figure 18]Cinnamein inhibits induction of NO production from LPS- and IFNγ-stimulated mouse RAW 264.7 macrophages. Figure 18A) Cells preincubated for 6 h with various concentrations of cinnamein were stimulated with 1 μg / ml LPS under serum-free conditions. After 24 h of stimulation, the level of nitrite in the supernatant was measured by Griess reagent. Figure 18B) Cells preincubated for various times with 400 μM cinnamein were stimulated with 1 μg / ml LPS under serum-free conditions. After 24 h of stimulation, the level of nitrite in the supernatant was measured. Figure 18C) Cells preincubated for 6 h with various concentrations of cinnamein were stimulated with 25 U / ml IFNγ under serum-free conditions. After 24 h of stimulation, the level of nitrite in the supernatant was measured. Results are the mean + SD of three independent experiments. *p<0.05, **p<0.01, ***p<0.001, NS, not significant. [Figure 19] Cinnamein inhibits LPS- and Poly IC-induced production of TNFα in primary mouse microglia. Microglia isolated from 2-day-old mouse pups were incubated with various concentrations of cinnamein for 6 h and subsequently stimulated with either 1 μg / ml LPS (FIG. 19A) or 50 μg / ml Poly IC (FIG. 19B) under serum-free conditions. After 24 h of stimulation, the levels of TNFα in the supernatants were measured by ELISA. Results are the mean + SD of three independent experiments. *p<0.05, **p<0.01, ***p<0.001, NS, not significant. [Figure 20] Cinnamein inhibits IL-1β production from LPS- and Poly IC-stimulated primary mouse microglia. Cells preincubated with various concentrations of cinnamein for 6 h were stimulated with either 1 μg / ml LPS (FIG. 20A) or 50 μg / ml Poly IC (FIG. 20B) under serum-free conditions. After 24 h of stimulation, the levels of IL-1β were measured in the supernatants by ELISA. Results are the mean + SD of three independent experiments. ***p<0.001. [Figure 21]Cinnamein reduces LPS- and Poly IC-induced production of IL-6 in primary mouse microglia. Microglia were incubated with various concentrations of cinnamein for 6 h and subsequently stimulated with either 1 μg / ml LPS (FIG. 21A) or 50 μg / ml Poly IC (FIG. 21B) under serum-free conditions. After 24 h of stimulation, the levels of IL-6 in the supernatants were measured by ELISA. Results are the mean + SD of three independent experiments. ***p<0.001. [Figure 22] Cinnamein inhibits the production of pro-inflammatory cytokines from Poly IC-stimulated primary mouse astrocytes. Astrocytes preincubated for 6 h with various concentrations of cinnamein were stimulated with 50 μg / ml of IC under serum-free conditions. After 24 h of stimulation, the levels of TNFα (Figure 22A) and IL-6 (Figure 22B) in the supernatants were measured by ELISA. Results are the mean + SD of three independent experiments. *p<0.05, **p<0.01, ***p<0.001. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0008] A.Definition "Nervous system injury", including central nervous system injury or peripheral nervous system injury, refers to any damage to the nervous system caused by trauma and / or disease.

[0009] The "central nervous system" (CNS) includes the brain, spinal cord, visual system, olfactory system, and auditory system. The CNS contains both neurons and glial cells, which are support cells that aid in the function of neurons. Oligodendrocytes, astrocytes, and microglia are glial cells within the CNS. Oligodendrocytes myelinate axons in the CNS, while astrocytes contribute to the blood-brain barrier, isolating the CNS from blood proteins and cells, and performing many supportive functions for neurons. Microglial cells perform immune system functions.

[0010] "Central nervous system injury" refers to injury to the central nervous system caused by trauma rather than disease. The term encompasses injury to the central nervous system that results in loss or impairment of motor function, sensory function, or a combination thereof.

[0011] The "peripheral nervous system" (PNS) includes the cranial nerves (except the optic and olfactory nerves) arising from the brain, the spinal nerves arising from the spinal cord, and the sensory neuronal cell bodies and their processes, i.e., all nervous tissue outside the CNS. The PNS includes both neurons and glial cells, which are support cells that aid in the function of neurons. Glial cells within the PNS are known as Schwann cells and are responsible for myelination of axons by providing a sheath that surrounds the axons.

[0012] "Peripheral nervous system injury" refers to injury to peripheral nerves caused by trauma, not disease. This term includes all degrees of nerve injury, including the lowest degree of nerve injury, known as neurapraxia, where the nerve remains intact but its signaling capacity is damaged. This term also includes a second degree, known as axotomy, where the axon is damaged but the surrounding connecting tissue remains intact. Finally, this term includes the last degree, known as neurotmesis, where both the axon and the connecting tissue are damaged.

[0013] "Traumatic brain injury" or "TBI" refers to an acquired brain or head injury in which the trauma causes damage to the brain. The damage can be localized, that is, limited to one area of ​​the brain, or diffuse, affecting one or more areas of the brain.

[0014] "Spinal cord injury" means damage to the spinal cord caused by trauma rather than disease. Depending on where the spinal cord and nerve roots are damaged, symptoms can vary widely, for example, from pain to paralysis to incontinence. Spinal cord injuries are described in various levels of "incomplete," which can vary from not affecting the subject to "complete" damage, meaning total loss of function. Spinal cord injuries have many causes, but are typically associated with significant trauma from car accidents, falls, sports injuries, and violence. The abbreviation "SCI" ​​stands for spinal cord injury.

[0015] "Spinal cord contusion" refers to injury caused by trauma rather than disease in which a portion of the spinal cord is crushed and injured, sparing some of its tissue, particularly the ventral nerve fibers that connect the cranial and caudal ends of the spinal cord.

[0016] "Nerve crush injury" refers to traumatic compression of a nerve by a blunt object such as a bat, surgical clamp, or other crushing object that does not result in complete severing of the nerve.

[0017] "Administering" refers to any method used to deliver a compound, salt, or composition to a subject. These include oral routes, intraduodenal routes, parenteral injection (including intravenous, subcutaneous, intraperitoneal, intramuscular, intravascular, or infusion), topical, and rectal administration. Those skilled in the art are familiar with administration techniques that can be used, for example, as discussed in Goodman and Gilman, The Pharmacological Basis of Therapeutics, current ed.; Pergamon, and Remington's, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, Pa. In some embodiments, the compounds and compositions are administered orally.

[0018] "Effective amount" refers to an amount of at least one administered agent or compound sufficient to alleviate or prevent to some extent one or more of the symptoms of the injury being treated. The result may be a reduction and / or alleviation of the signs, symptoms, or causes of the injury, or any other desired change in a biological system. For example, an "effective amount" for therapeutic use is the amount of compound required to cause a clinically significant reduction in the progression or severity of the symptoms associated with the injury being treated. The appropriate "effective" amount in any individual case may be determined using techniques such as a dose escalation study.

[0019] A "subject" can be any living subject, including a mammalian subject, such as a human.

[0020] "Prodrug" refers to any pharma- ceutically acceptable compound or salt that, upon administration to a subject, can yield a benzoate, either directly or indirectly, e.g., by metabolism in the body.

[0021] "Pharmaceutically acceptable" refers to a substance such as a carrier, diluent, or excipient that does not abolish the biological activity or properties of the active ingredients and that is relatively non-toxic, i.e., the material may be administered to a subject without causing undesired biological effects or interacting in a deleterious way with any of the components of the composition in which it is contained.

[0022] A "pharmaceutical composition" refers to a composition comprising a biologically active compound, optionally mixed with at least one pharma- ceutically acceptable component, such as a carrier, stabilizer, diluent, dispersing agent, suspending agent, thickening agent, or excipient.

[0023] B. Treatment method In one aspect, a method of slowing the progression or reducing the severity of symptoms associated with nervous system injury in a subject in need thereof comprises administering to the subject an effective amount of a benzoate salt or a prodrug thereof, thereby slowing the progression or reducing the severity of symptoms associated with nervous system injury.

[0024] In one aspect, the benzoate salt, if used, is sodium benzoate, potassium benzoate, calcium benzoate, 2-aminobenzoic acid, 3-aminobenzoic acid, 4-aminobenzoic acid, or any combination thereof.

[0025] In one aspect, the prodrug of benzoate, if used, is benzyl cinnamate, glyceryl tribenzoate, cinnamic acid, benzyl acetate, benzyl alcohol, benzoic acid, quinic acid, phenylalanine, tyrosine, or any combination thereof.

[0026] Various nervous system disorders can be treated using the disclosed methods. In one embodiment, the nervous system injury in the subject is a central nervous system (CNS) injury or a peripheral nerve injury. In a further embodiment, the nervous system injury is a spinal cord injury (SCI), a spinal cord contusion, or a nerve crush injury. For example, when the injury to the nervous system is a spinal cord injury (SCI), benzoate or a prodrug thereof can improve the nervous system dysfunction caused by trauma to the cervical, thoracic, lumbar, or sacral region of the spinal cord, including but not limited to the dysfunction caused by trauma to one or more of the following dermatomes: C1, C2, C3, C4, C5, C6, C7, T1, T2, T3, T4, T5, T6, T7, T8, T9, T10, T11, T12, L1, L2, L3, L4, or L5.

[0027] In one embodiment, the nervous system injury is traumatic brain injury (TBI). In various embodiments, the TBI can be injury to the frontal lobe, parietal lobe, occipital lobe, temporal lobe, brain stem, or cerebellum. In some embodiments, the TBI is mild TBI. In further embodiments, the TBI is moderate to severe TBI. Benzoate and its prodrugs can, in various embodiments, cause detectable improvement or reduced progression of one or more of the following symptoms of TBI: headache, memory problems, attention deficit, mood swings and frustration, fatigue, visual impairment, memory loss, reduced attention or concentration, sleep disorders, dizziness or loss of balance, irritability, emotional disturbance, depression, seizures, nausea, loss of smell, sensitivity to light and sound, mood changes, confusion or confusion, or slow thinking.

[0028] In another embodiment, the nervous system injury is a demyelinating disorder, which can be, for example, optic neuritis, X-adrenoleukodystrophy, Krabbe disease, progressive multifocal leukoencephalopathy, adrenomeleoneuropathy, acute disseminated encephalomyelitis, acute hemorrhagic leukoencephalitis, multiple sclerosis, Balo's disease (concentric sclerosis), Charcot-Marie-Tooth disease, Guillain-Barré syndrome, HTLV-I associated myelopathy, neuromyelitis optica (Devic's disease), Schilder's disease, transverse myelitis, or a combination thereof.

[0029] In general, the damage that can be treated by the disclosed method can result in some symptoms that can be alleviated, delayed, or prevented using benzoate or its prodrug.In one aspect, administering an effective amount of benzoate or its prodrug results in a reduction in glial inflammation, an improvement in motor function or coordination, or an improvement in learning or memory dysfunction.In a further aspect, particularly when the damage to be treated is damage to the CNS, administering an effective amount of benzoate or its prodrug prevents or reduces the severity of symptoms related to mental depression.A non-limiting example of a symptom of mental depression is the level of physical activity that a subject is motivated to engage in.

[0030] In one embodiment, it may be useful to administer benzoate or a prodrug thereof before nervous system damage has progressed significantly. For example, an effective amount of benzoate or a prodrug thereof may be administered within 24 hours after nervous system damage, for example, within 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 hour after nervous system damage. In another embodiment, an effective amount of benzoate or a prodrug thereof may be administered 24 hours or more after nervous system damage. In a further embodiment, an effective amount of benzoate or a prodrug thereof may be administered within 24 hours after nervous system damage, and administration of benzoate or a prodrug thereof may continue for a period of time, for example, days, weeks, months, or years after injury.

[0031] In one embodiment, the subject is diagnosed with a nervous system injury prior to the administering step. In a further embodiment, the subject is diagnosed with a central nervous system (CNS) injury or a peripheral nerve injury prior to the administering step. In a further embodiment, the subject is diagnosed with a spinal cord injury (SCI), spinal cord contusion, or nerve crush injury prior to the administering step. In yet a further embodiment, the subject is diagnosed with a traumatic brain injury (TBI) prior to the administering step.

[0032] In one aspect, prior to the administering step, the subject being treated has not been diagnosed with or been treated for a urea cycle disorder, glycine encephalopathy, multiple sclerosis, Parkinson's disease, Alzheimer's disease, Huntington's disease, or an autism spectrum disorder, examples of which include Asperger's syndrome, childhood disintegrative disorder, and pervasive developmental disorder.

[0033] In one embodiment, the subject being treated is over 12 years of age. In a further embodiment, the subject being treated is at least 18 years of age. In yet another embodiment, the subject being treated is at least 21 years of age. In other embodiments, the subject can be of any age, including under 12 years of age.

[0034] In one embodiment, the benzoate or prodrug thereof may be administered as a pharmaceutical composition comprising the benzoate or prodrug thereof and a pharma- ceutically acceptable excipient, the composition comprising more than 0.1% benzoate or prodrug thereof by weight of the composition, e.g., more than 0.5%, more than 1%, more than 2%, more than 5%, more than 10%, more than 15%, more than 20%, more than 30%, more than 40%, or more than 50% benzoate or prodrug thereof by weight based on the total weight of the pharmaceutical composition, up to 99% benzoate or prodrug thereof by weight. For example, the pharmaceutical composition may comprise benzoate or prodrug thereof in an amount ranging from 1.1% to 50% or more by weight of the composition.

[0035] In a further aspect, the pharmaceutical composition may comprise only benzoate or a prodrug thereof as an active ingredient for treating injury. In other words, in one aspect, benzoate or a prodrug thereof may serve as the sole active ingredient. In a further aspect, the pharmaceutical composition of the present disclosure comprises an active pharmaceutical ingredient that consists essentially of, or in other aspects consists of, benzoate or a prodrug thereof.

[0036] In one embodiment, the total daily dose of benzoate or a prodrug thereof is 100 mg / day, 120 mg / day, 150 mg / day, 180 mg / day, 200 mg / day, 225 mg / day, 250 mg / day, 300 mg / day, 400 mg / day, 500 mg / day, 1000 mg / day, 1500 mg / day, 2000 mg / day, 2500 mg / day, 3000 mg / day, 3500 mg / day, or 4000 mg / day, administered to a subject as a single dose or multiple doses, depending on various factors such as the route of administration. In a further embodiment, the total daily dose of benzoate or a prodrug thereof is 1000 mg / day to 4000 mg / day, e.g., 1000 mg / day to 3000 mg / day, or 1000 mg / day to 2000 mg / day.

[0037] In one particular embodiment, these total daily doses of benzoate or its prodrugs can be orally administered as a single dose or multiple doses.In one embodiment, the composition administered to the subject can be formulated in a form suitable for oral administration.For example, the composition can be formulated in the form of dry powder, tablet, lozenge, capsule, granule, or pill.Pharmaceutically acceptable excipients include, but are not limited to, fillers, binders, preservatives, disintegrants, lubricants, suspending agents, wetting agents, solvents, surfactants, acids, flavoring agents, polyethylene glycol (PEG), alkylene glycol, sebacic acid, dimethyl sulfoxide, alcohol, or any combination thereof. EXAMPLES

[0038] C. Working Example The following examples further illustrate the present disclosure. The scope of the disclosure and claims are not limited by the scope of the following examples.

[0039] 1. Materials and Methods A. Animals Male C57BL6 mice (7-8 weeks old) purchased from Harlan, Indianapolis, IN were used in this study. Animal maintenance and surgical procedures were performed in accordance with the NIH guidelines of the Care and Use Committee and approved by the Rush University Animal Care and Use Committee. Animals were housed in an environment with a stable temperature and a 12-h light / dark cycle. Water and food were provided ad libitum.

[0040] b. Controlled Cortical Impact Procedure To induce brain injury in mice, the controlled cortical impact (CCI) injury technique was applied. Adult C57BL6 mice were anesthetized with 2% isoflurane and allowed to breathe normally without tracheal intubation. Body temperature was maintained at 37 °C on a heating pad during the surgical procedure and monitored by a rectal probe. The depth of anesthesia was observed by gently pinching the toes without causing any injury. The head of the anesthetized mouse was shaved with a sterile electric shaver and the skin was washed with betadine solution. The animal's head was then fixed in a stereotaxic frame and TBI was induced using the CCI technique (Figure 1A-1D). First, a midline skin incision was performed to expose the skull, and a stereotaxic device was used to make a 4 mm diameter craniotomy on the right side of the exposed skull at coordinates -1.5 mm AP and -1.5 mm ML. This burr hole then exposed the brain with the dura intact. Under surgical microscope control, a Leica Impact One Stereotaxic Impactor (Leica Mi-crosystems, Buffalo Grove, IL) fitted with a 1.0 mm rounded metal tip was tilted vertically towards the brain surface where the dura was intact. Subsequently, a mild injury was unilaterally induced on the right side of the exposed brain region with a strike speed of 1.0 m / s. A sterile sponge fixation board was used to support the underside of the head to act as a cushion-like support during the induction of brain injury. After the impact injury, damage occurred in the cerebral cortex, causing extensive structural damage in the surrounding area. Animals in the sham group underwent a similar surgical procedure but without CCI injury. The operated animals were then removed from the stereotaxic holder and the skin incision was lightly sutured to close the incision area. All operated animals were placed in a warming blanket to maintain body temperature within normal limits. These animals were monitored until recovery from anesthesia and for the next 3 consecutive days postoperatively.

[0041] Generating clinically relevant TBI models using small laboratory animals such as mice is a challenging task in TBI research. The impact of TBI may have varying physical and psychological outcomes depending on the extent of damage to the brain. Some symptoms may appear immediately after injury, while others may appear days or weeks later. Therefore, for standardization purposes, a fixed 1 mm rounded tip was used at various speeds. In this study, mice were randomly divided into three groups, and CCI was applied with a 1 mm rounded tip at three speeds, namely 1.0 V, 1.25 V, and 1.50 V, to induce mild, moderate, and severe injury, respectively (Figure 1C). At the end of one week after surgery, all three groups of animals were perfused with 4% paraformaldehyde and the brains were removed. Brain sections were then prepared at a thickness of 40 μm. Cresyl violet staining was used to study the histopathological features of brain damage, which revealed significant tissue damage in the cortical and hippocampal regions in the mild injury group. However, no significant damage was found in the contralateral side of the brain in this group of mice. In the moderate injury group, more damage was found in the tissues of the ipsilateral cortex and hippocampus regions of the mouse brain, and the recovery of the mice after surgery was very slow and in some cases fatal. Furthermore, in the severe injury group, severe tissue damage was found in both the ipsilateral cortex and hippocampus of the brain after surgery (Figure 1C). The recovery of the mice was minimal, and the damage produced often became fatal in this group of mice. Therefore, based on the histopathological observation of the three types of injury groups, a mild type of CCI injury (1 mm tip and 1.0 V) was selected to depict the beneficial effects of cinnamon metabolite NaB in improving cognitive and motor functions after brain injury (Figure 1C).

[0042] c. Treatment with sodium benzoate or sodium formate Sodium benzoate ("NaB") and sodium formate ("NaFO") were solubilized in 0.1% methylcellulose solution. Starting 24 hours after CCI injury, mice were orally treated with NaB or NaFO (50 mg / kg / day) once a day for 7 days after surgery. Oral treatment was then continued every other day until 21 days after surgery, and after behavioral analysis, mice were sacrificed for histological and biochemical studies.

[0043] d. Experimental groups and NaB / NaFO treatments The experimental design used in this study is shown in Figure 1D. All mice were randomized into the following groups:

[0044] Group 1: Control / sham group (n=6 per group): Mice underwent surgery without injury or treatment.

[0045] Group 2: CCI group (n=6 per group): mice received a CCI injury and no treatment was administered.

[0046] Group 3: CCI+NaB treatment (n=6 per group): mice were subjected to CCI and oral NaB (50 mg / kg / day) treatment was started 24 hours after the induction of injury.

[0047] Group 4: CCI+NaFO treatment (n=6 per group). Mice were subjected to brain injury and oral NaFO (50 mg / kg / day) treatment was started 24 hours after induction of injury.

[0048] e. Western blotting Western blotting was performed as described in a previous study. Equal amounts of protein were electrophoresed in 10% or 12% SDS-PAGE and transferred onto nitrocellulose membranes. Blots were probed with primary antibodies overnight at 4°C. The following are the primary antibodies used in this study and are detailed in Table 1 below: anti-iNOS (1:1000, BD Bio-sciences), anti-Iba1 (1:1000, Abcam), anti-GFAP (1:1000, Santa Cruz Biotechnology, Dallas, TX), and anti-β-actin (1:5000, Abcam). After overnight incubation, the primary antibodies were removed, the blots were washed with phosphate buffered saline containing 0.1% Tween-20 (PBST), and the corresponding infrared fluorophore-tagged secondary antibodies (1:10,000, Jackson Immuno-Research) were added at room temperature. The blots were then incubated with the secondary antibodies for 1 hour. Blots were then scanned with an Odyssey infrared scanner (Li-COR, Lincoln, NE), and band intensities were quantified using ImageJ software (NIH, USA). [Table 1]

[0049] f. Immunohistochemistry Mice were anesthetized with ketamine-xylazine mixed solution and perfused with PBS, followed by 4% paraformaldehyde (w / v) in PBS, followed by dissection of the brains for immunofluorescence microscopy. Briefly, dissected brains were incubated in 10% sucrose for 3 h, followed by 30% sucrose overnight at 4 °C. Brains were then embedded in optimal cutting temperature medium (Tissue Tech) at -80 °C and processed for conventional cryosectioning. Frozen sections (40 μm thick) were treated with chilled ethanol (-20 °C), washed with PBS, blocked with 2% BSA in PBST, and double-labeled with two primary antibodies (Table 1). After three washes with PBST, sections were incubated with Cy2 and Cy5 (Jackson ImmunoResearch Laboratories). Sections were mounted and observed under an Olympus IX81 fluorescent microscope. Counting analysis was performed using Olympus Microsuite V software with the aid of a touch counting module.

[0050] g. Quantification of Lesion Volume Using Stereological Techniques Lesion volume estimation was performed based on the Cavalieri method of unbiased stereology using Stereo Investigator software (MicroBright Biosciences, USA). The Cavalieri estimator with a grid spacing of 1 mm was used to determine the volumes of both the ipsilateral and contralateral hemispheres of the brain. Every fourth section was analyzed starting from a random starting point. Lesion volume was estimated by subtracting the volume of the ipsilateral hemisphere from the volume of the contralateral hemisphere. The volume of the lesion cavity estimated in brain sections of untreated mice was then compared to the lesion volume in brain sections of drug-treated mice.

[0051] h. Behavior analysis Behavioral analysis of animals was performed on post-surgery days 7 and 21 after CCI injury. These time points for behavioral testing were selected based on previous studies using these animal models in which behavioral abnormalities were observed at these time points.

[0052] i. Open field behavior The animals' performance in the open field test was analyzed as described in a previous study. Briefly, each animal was allowed to move freely for 5 min to explore an open field activity area with a wooden floor in the shape of a square measuring 40x40 cm and with walls 30 cm high. A video computer 6 (Basler Gen I Cam-Basler acA 1300-60) connected to a Noldus computer system was fixed facing downwards on the top above the open field activity area. Each mouse was placed individually in the center of the activity area and the performance was monitored by a live video tracking system. The central area was arbitrarily defined as a square of 20x20 cm (half of the total area).

[0053] j. Rotarod The coordination and balance of the animals' forelimb movements were observed using the rotarod test, as described in a previous study. Briefly, each mouse was placed on a confined section of the rod, and the test began with a smoothly increasing speed from 4 rpm to 40 rpm for 5 min. If the mouse did not fall off the rod, it was removed from the rod after 5 min. Fall latency was measured in seconds and used for analysis. After CCI injury, each mouse performed the task in three trials during the test session, and the average score of these three trials was used as the individual rotarod score. A test on each rod was terminated if the mouse fell off the rod or completed an inappropriate rotation while hanging and held on the rod.

[0054] k. Tail suspension test Mice were subjected to the tail suspension test using the methodology described in a previous study. Mice were gently suspended upside down by their tails 50 cm above the floor using non-toxic adhesive tape for 6 min. Immobility time was defined as the period during which the mouse was only passively hanging without active movement. Increased immobility time was defined as depression-like behavior.

[0055] l. Nest-building behavior Nests consisting of 5 cm x 5 cm compressed cotton squares were placed in the cages between 5:00 PM and 6:00 PM. The following morning between 9:00 AM and 10:00 AM, two observers blinded to the experimental procedures scored the quality of the nests constructed by the mice using a 5-point scale as follows: score 1 (>90% nests intact), score 2 (50%-90% nests intact), score 3 (10%-50% nests intact but with no recognizable nest location), score 4 (<10% nests intact and nests are recognizable but flat), score 5 (<10% nests intact and nests are recognizable with walls higher than the mouse body).

[0056] m.Beam Runway The beam runway was made of smooth wood and measured 65 cm long x 0.7 cm wide x 4 cm high. A black box with an opening was fixed at one end and an aversive stimulus (a bright lamp) was fixed at the other end of the beam. This test was used to evaluate the complex coordination and balance of mice while crossing the beam, and the procedure was performed as described in a previous study. The mouse was placed on the beam near the light source and the light was turned "on". This forces the animal to enter the box to avoid the aversive stimulus, which is then turned off. Six repetitions were performed with a 2-minute rest period inside the box. The parameters measured were the time taken (in seconds) to reach the box and the number of steps with dragging / slipping of the contralateral limb. Errors were taken into account whenever the number of slips and slides of the foot on the beam were counted. Beam gait analysis was performed by an observer blinded to the treatment on the 7th and 21st days after surgery.

[0057] n. Lattice runway A grid runway (65 cm long x 8 cm wide x 1 cm spacing) made of parallel grid bars with 1 cm spacing between bars was kept on the table surface during the testing session. A protective soft pad was placed under the grid runway to avoid serious injury if the animal fell off the grid. Each mouse was allowed to walk freely on the grid, and the time and number of steps taken to traverse the runway were recorded. Each successful foot placement on the grid was recorded as a step. However, an error was considered if the foot slipped through the grid or if the foot missed the bar and extended downward through the plane of the bar. The locomotor behavior of the animals on the grid was assessed by an observer blinded to the treatment on days 7 and 21 after CCI injury.

[0058] o. Barnes maze test The Barnes maze test was performed as described in previous studies [44,49,58]. Briefly, mice were first trained for 2 consecutive days and then tested on the third day. After each training session, the maze and the escape tunnel were thoroughly cleaned with mild detergent to avoid instinctive odor avoidance due to the mouse's odor from familiar objects. On the third day, a video camera (Basler Gen I Cam-Basler acA 1300-60) connected to a Noldus computer system was placed above the maze and illuminated with a high-voltage light that produced enough light and heat to motivate the animals to enter the escape tunnel. Performance was monitored by a video tracking system (Noldus System). Cognitive behavioral parameters were investigated by measuring the latency (the period before all four paws were on the floor of the escape box) and errors (incorrect responses before all four paws were on the floor of the escape box).

[0059] pT-shaped maze Mice were first habituated to the T-maze under food-deprived conditions for 2 days. Food rewards were provided at least five times during 10 min of training. The T-maze was cleaned with a mild detergent solution between each test session to minimize the ability of the animals to use any olfactory cues. The side of the food reward was always associated with a visual cue. Every time an animal consumed a food reward, it was considered a positive turn.

[0060] q. Novel Object Recognition (NOR) Test This test evaluates the animal's ability to recognize novel objects in the environment and monitor short-term memory. First, mice were placed in a square novel box (20 inches long x 8 inches high) surrounded by infrared sensors. Two plastic toys (2.5-3 inches in size) differing in color, shape, and texture were placed in specific locations in the environment, 18 inches apart from each other. Mice were allowed to freely explore the environment and objects for 15 minutes and then returned to their individual home cages. After a 30-minute interval, mice were returned to the environment with the two objects in the same location, but this time one of the familiar objects was replaced by a third novel object. Mice were again allowed to freely explore both objects for 15 minutes. The familiar and novel objects were thoroughly washed with mild detergent after each test session.

[0061] r.Statistical analysis Based on previous studies of similar type and complexity, six mice are expected to perform >80% of the effort for all behavioral experiments. Statistical analysis was performed using Student's t-test for two-group comparisons and, when appropriate for multiple comparisons, one-way ANOVA followed by Tukey's multiple comparison test by using GraphPad Prism7. Data are presented as mean ± SD or mean ± SEM as per the indicated figure legends. Statistical significance was determined at the p<0.05 level.

[0062] 2.NaB treatment attenuates glial activation in CCI-induced TBI mice Recent findings have established microglial and astroglial activation and associated neuroinflammation as key pathological events in various neuroinflammatory and neurodegenerative disorders, including brain injury. Shortly after the initial CCI injury, the tissue environment is modified to activate glial cells. Accordingly, after CCI injury (Figure 1), a marked increase in the number of GFAP-positive astrocytes (Figures 2A, 2B, 2E, and 2F) and Iba1-positive microglia (Figures 3A, 3B, 3E, and 3F) was observed in the cortical and hippocampal regions of mice 7 days after injury compared to sham controls. Western blot analysis of hippocampal extracts also confirmed this increase in GFAP (Figures 2I and 2J) and Iba1 (Figures 3I and 3J). However, oral treatment of CCI-injured TBI mice with NaB resulted in a decrease in both GFAP-positive astrocytes (Figures 2A-2F) and Iba1-positive microglia (Figures 3A-3F). This result was specific, as sodium formate (NaFO) remained unable to inhibit glial activation in the hippocampus following good TBI (Figures 2A, 2B, 2E, and 2F and Figures 3A, 3B, 3E, and 3F).

[0063] Decreased and / or normalized levels of GFAP (Figures 2I-2J) and Iba1 (Figures 3I-3J) proteins in the hippocampus of TBI mice treated with NaB are also evident from Western blots. Activated glial cells are known to express inducible nitric oxide synthase (iNOS), which produces excess nitric oxide to cause nitrite stress in a neuroinflammatory environment. Correspondingly, levels of iNOS were higher in the cortex and hippocampus of TBI mice 7 days after injury compared to sham controls (Figures 2 and 3). Double-labeling immunofluorescence analysis revealed that increased iNOS was present in both GFAP-expressing astrocytes (Figures 2A-2H) and Iba1-positive microglia (Figures 3A-3F). However, treatment of TBI mice with NaB, but not NaFO, led to inhibition of iNOS in both the cortex and hippocampus (Figures 2C-2D and 3C-3D). These findings were confirmed by quantitative analysis (Figures 2G-2H) and Western blot (Figures 3I-3J). Collectively, these results indicate that NaB can reduce glial inflammation in vivo in the CNS of CCI-induced TBI mice.

[0064] 3. Oral NaB stimulated remyelination in mice with TBI Proteolipid protein (PLP) is a marker for oligodendrocytes, and A2B5 is a marker for oligodendrocyte precursor cells (OPCs). After 21 days of treatment, brain sections were double-labeled with antibodies against PLP and A2B5. The images in Figure 4 show that, as expected, the level of PLP was very low in the corpus callosum of mice with TBI, and many A2B5-positive OPCs were localized in the demyelinated areas. However, NaB treatment significantly increased the level of PLP in the corpus callosum of mice with TBI. Thus, NaB treatment also reduced the number of OPCs in the corpus callosum of TBI mice. These results are specific to NaB, and NaFO, a molecule structurally similar to NaB without a benzene ring, remained unable to restore the level of PLP and reduce the number of OPCs in the corpus callosum of TBI mice.

[0065] 4. NaB treatment reduced lesion volume in CCI-induced mice Since oral NaB reduced glial inflammation in the CNS of TBI mice, we then investigated whether NaB treatment could reduce the lesion volume. Therefore, lesion volumes were measured in cresyl violet stained sections and compared between untreated and treated groups. Figure 5A shows cresyl violet stained brain sections arranged serially to demonstrate the volume of the lesion cavity from the different mouse groups. 21 days after injury, typical lesions including enlarged cavities were observed in CCI-induced TBI mice, starting from the cortex and connecting to the lateral ventricle via the hippocampus, compared to none observed in sham controls (Figure 5B). On the other hand, oral administration of NaB, but not NaFO, reduced the size of the lesion cavity in CCI-induced mice. Quantitative analysis of lesion volumes using Cavalieri stereological techniques revealed that all lesion volumes in all hemispheres were significantly reduced after oral treatment of NaB when compared to either untreated or NaFO-treated TBI mice (Figure 5C).

[0066] 5. Oral NaB improved open-field behavior and locomotor activity in mice with TBI The most important therapeutic objective of neuroprotection studies is to limit secondary tissue loss and maintain or improve behavioral function. Therefore, to analyze whether oral administration of NaB protects not only the tissue damage but also the functional deficit caused by CCI injury, the overall locomotor activity was investigated. A video camera 6 (Basler Gen I Cam-Basler acA 1300-60) connected to a Noldus computer system remained mounted facing downwards on the top over the open field activity arena to record general locomotor behavior. Figure 6A and Figure 6F show heat maps summarizing the overall activity of mice in the open field test 7 and 21 days after injury, respectively. Compared to either untreated or NaFO-treated TBI mice, general locomotor activity showed significant improvement (Figure 6A-6E) in TBI mice treated with NaB 7 days after injury. Functional improvement was clearly visualized from the distance traveled (Figure 6B), speed (Figure 6C), center frequency (Figure 6D), and rearing behavior (Figure 6E). On the other hand, no significant differences in overall movement were observed between treated and untreated TBI groups 21 days after injury (Figure 6).

[0067] Subsequently, the recovery of motor coordination and balance activity in all groups of CCI-injured mice was also investigated using the rotarod test 7 and 21 days after injury. After CCI injury, mice without treatment showed a significant decrease in the latency to fall 7 days after injury, and this motor activity remained impaired on the rotarod for 21 days after injury compared to the sham control group. However, treatment of CCI-injured mice with NaB, but not NaFO, resulted in a prolonged latency by maintaining proper body movement and balance function on the rotarod test (Figure 6L).

[0068] Depression is a common symptom found during the early stages of brain injury. Therefore, we monitored depression-like behavior in CCI-injured mice. Previous studies in TBI research have demonstrated that depression in mice can be analyzed by increased immobility periods. Therefore, this study was performed to investigate the neuroprotective effect of NaB on depression-like behavior in CCI-injured mice. Seven days after injury, CCI mice without any treatment showed significantly longer immobility times than sham controls (Figure 6K). On the other hand, CCI mice treated with NaB showed significantly less immobility times compared to either untreated or NaFO-treated mice. Upon NaB treatment, the immobility periods were close to normal levels. These results suggest that NaB can control depression-like behavior in CCI-injured mice.

[0069] TBI-induced damage always impairs the connection between the brain and muscles, ultimately affecting locomotion. Therefore, the gait-related impairments of CCI mice on the beam and grid were analyzed because these two multi-sided runways appeared to reveal different patterns of movement than those on the open-field behavioral test. Previous studies have revealed that these beam and grid runways are particularly useful in models of unilateral TBI, as they give scientists the opportunity to analyze and compare the movements of the contralateral versus ipsilateral limbs. Therefore, we investigated the neuroprotective role of NaB on the recovery of gait function in a unilateral CCI model using the beam and grid runway. CCI mice tended to drag the contralateral pelvic limb while walking. This type of behavior was not seen in the sham controls. Moreover, the sham controls did not show any significant changes in the latency to cross the beam or leg steps after surgery.

[0070] However, none of the CCI mice were able to cross the beam on the day of surgery and the day after surgery (Figures 6M-6O). Seven days after injury, CCI mice without treatment showed significant deficits in balancing the body on the beam or slipped their feet to pass the grid. CCI mice without treatment showed a reduced ability of locomotor behavior, showing more latency, step and leg defects, or misplacement of legs while crossing the beam compared to sham controls. Similar results were seen for the grid analysis (Figures 6P-6R). However, when treated with NaB but not NaFO, CCI-injured mice showed a significant improvement in locomotor movement on the beam and grid runway.

[0071] NaB-treated CCI mice also showed significant improvements in latency, leg step, leg slip, and leg misplacement compared to either untreated or NaFO-treated CCI mice (Figures 6M-6R). On the other hand, 21 days after injury, CCI mice were significantly restored to near-normal levels, such that no significant changes in these parameters were observed with respect to sham controls. As a result, NaB treatment also did not show significant protection in either beam walking or grid runway walking in CCI mice 21 days after injury.

[0072] 6. Oral NaB protected spatial learning and memory in mice with TBI TBI survivors often suffer from lifelong learning and memory problems. Therefore, to investigate whether oral NaB protects memory and cognitive function in TBI mice, we monitored the mice's performance on novel object recognition (NOR), Barnes maze, and masked T-maze. Figure 7A shows a heatmap depicting the novel object recognition of mice after 21 days of treatment. Figure 7C shows the exploration time results of this same test.

[0073] The Barnes circular maze test is a hippocampus-dependent cognitive task that requires spatial reference memory. Figure 7B shows the heat map of the results of the Barnes circular test of TBI mice 21 days after treatment, Figure 7D shows the latency, and Figure 7E shows the number of errors made. TBI mice do not easily find the reward hole, need more time (latency) and make more errors. On the other hand, TBI mice treated with NaB have the same ability as healthy control mice in finding the target hole with less latency and fewer errors.

[0074] Similar results were found in the T-maze test. Figure 5F shows the number of positive turns and Figure 5G shows the number of negative turns made during this test after 21 days of treatment. TBI mice made fewer positive turns and more negative turns than sham controls. Again, NaB treatment significantly improved hippocampus-dependent memory performance in TBI mice, as indicated by a higher number of positive turns and a lower number of negative turns.

[0075] These results were specific to NaB: its negative control, NaFO, remained unable to improve hippocampal-dependent behaviors in TBI mice.

[0076] 7. Discussion TBI is the leading cause of death and disability in the United States, yet despite extensive investigations, no effective treatment is available to date to improve the quality of life of TBI patients, except for regular medical evaluation and care. Thus, describing safe and effective treatments to modulate the pathological process of TBI and result in improved behavioral outcomes is an important area of ​​research. Several pieces of evidence outlined in this study clearly support the conclusion that NaB can inhibit the disease process of TBI in a CCI-induced mouse model. Although TBI caused large lesion cavities, oral NaB treatment starting 24 h after CCI reduced the lesion volume and restored the integrity of the structural organization of the damaged hippocampus. In contrast, treatment with NaFO, a NaB analog without a benzene ring, remained unable to show such protection. NaB treatment also reduced depression-like behavior, attenuated motor dysfunction, and enhanced cognitive performance in mice with TBI. Moreover, oral NaB did not cause any side effects (e.g., weight loss, hair loss, fecal bolus, infections, inappropriate behavior, etc.), consistent with its safety track record. These results suggest that oral NaB may be beneficial in the treatment of TBI and that NaB should not be toxic to TBI patients.

[0077] Glial activation and upregulation of pro-inflammatory molecules in the CNS are involved in the pathogenesis of several neurodegenerative diseases, including TBI. It is known that immediately after TBI, microglia and astroglia in the brain are activated and produce toxic amounts of pro-inflammatory cytokines (e.g., IL-1β, TNFα, etc.), pro-inflammatory enzymes (e.g., inducible nitric oxide synthase or iNOS), reactive oxygen species, etc. for a long period of time, ultimately causing axonal damage. Here, it is shown that NaB treatment reduces the levels of microglial marker Iba1 and astroglial marker GFAP in the hippocampus of mice with TBI, and decreases the expression of iNOS. Thus, NaB treatment, started 24 hours after TBI in a therapeutic mode, can reduce and / or normalize glial inflammation in TBI mice.

[0078] The signaling mechanisms by which glial cells are activated are poorly understood. NaB has been reported to inhibit LPS-induced expression of iNOS and proinflammatory cytokines in microglia. TLR4 is the prototype receptor for LPS. However, NaB does not affect the levels of TLR4 in LPS-stimulated microglia, indicating that NaB prevents LPS-induced expression of proinflammatory molecules without engaging its receptor TLR4. Interestingly, it is the intermediates (HMG-CoA, mevalonate, and farnesyl pyrophosphate) of the mevalonate pathway, but not the end products (cholesterol and coenzyme Q), that reverse the anti-inflammatory effects of NaB in microglia. Suppression of LPS-induced activation of NF-κB and expression of iNOS in glial cells by farnesyltransferase inhibitors presents a critical role for the farnesylation reaction in upregulating the iNOS gene. Consistent with the role of farnesylation in the activation of p21ras, it is found that p21ras signaling plays a key role in the expression of proinflammatory molecules in glial cells. Therefore, inhibition of p21ras activation in microglial cells by NaB indicates that NaB attenuates glial inflammation through inhibition of p21ras activation.

[0079] So far, there is no effective inhibitory therapy available to stop the progression of TBI. Anticoagulants exist to prevent blood clots and improve blood flow, but anxiolytics to reduce fear and nervousness, antidepressants to treat symptoms of depression and mood lability, anticonvulsants to prevent seizures, and muscle relaxants to reduce muscle spasms are peripheral treatments, except for other anticoagulants. Furthermore, some of these drugs show limited symptom relief with some side effects. On the other hand, there are several advantages of NaB over available TBI therapies. First, NaB is objectively safe. It is water-soluble and, when consumed in excess, is secreted through urine. Second, NaB can be taken orally, which is the least painful route of drug treatment. Oral NaB reduced glial activation in vivo in the hippocampus and improved cognitive performance in TBI mice. Third, NaB is economical compared to other existing anti-TBI therapies. Fourth, the entry of drugs through the blood-brain barrier (BBB) ​​is a key issue for the treatment of CNS disorders. In the early stages of TBI, the BBB remains compromised, but over time, the integrity of the BBB improves, and thus BBB-permeable drugs would be useful for neuroprotection in TBI patients. NaB has also been detected in the brains of mice orally treated with cinnamon. Thus, after oral treatment, NaB enters the brain.

[0080] 8. Protection of Mice from Controlled Cortical Impact Injury by the Food Additive Glyceryl Tribenzoate Here, we investigated the neuroprotective effects of GTB in a controlled cortical impact (CCI) mouse model of TBI. We show that after oral administration, GTB can attenuate glial activation, reduce the levels of pro-inflammatory molecules, decrease lesion volume, and improve synaptic structure in CCI-induced TBI mice. Functionally, oral GTB restored motor skills and improved learning and memory in TBI mice, highlighting the possible therapeutic application of GTB in TBI.

[0081] 9. Oral GTB attenuates astroglial and microglial activation in CCI-induced TBI mice Astrocytes and microglia are two important cell types in the central nervous system, however, research over the past three decades has revealed that upon activation, these cells release distinct proinflammatory molecules and are involved in the pathogenesis of various neuroinflammatory and neurodegenerative disorders, including TBI.

[0082] Therefore, we investigated the effect of oral GTB on glial activation in the CNS of TBI mice. First, we monitored astroglial activation, and as expected, CCI injury induced astroglial activation in the cortex and hippocampus, as revealed by enhanced GFAP expression 7 days after injury compared to sham controls (Figures 8A-8B). This finding was supported by counting GFAP-positive cells in both the cortex (Figure 8E) and hippocampus (Figure 8F). The increase in GFAP after TBI was further confirmed by Western blot analysis of hippocampal extracts (Figures 8I-8J). Recently, we found that oral administration of GTB at a dose of 50 mg / kg body weight / day alleviated Huntington's pathology in mice and inhibited adoptive transfer of experimental allergic encephalomyelitis (EAE) in mice, an animal model of multiple sclerosis (MS). Thus, here, we treated CCI-injured mice with GTB at a dose of 50 mg / kg body weight / day by oral gavage and observed a decrease in GFAP-positive astrocytes (Figures 8A-8F) and GFAP protein levels (Figures 8I-8J) in the hippocampus of TBI mice upon GTB treatment. This result was specific, as we found no such changes upon vehicle treatment (Figures 8A-8F and 8I-8J). Activated astrocytes express various pro-inflammatory molecules, including inducible nitric oxide synthase (iNOS), which is known to produce excess nitric oxide to cause nitrite stress in a neuroinflammatory environment.

[0083] Therefore, we investigated the status of iNOS in the hippocampus and cortex of GTB-treated and untreated TBI mice. As expected, we also found an increase in iNOS-positive cells (Figures 8A, 8B, 8G, and 8H) and iNOS protein levels (Figures 8K-8L) in the brains of TBI mice compared with sham controls. Many GFAP-positive astrocytes colocalized with iNOS (Figures 8A-8D). However, similar to the inhibition of astroglial activation, oral GTB also decreased iNOS-positive cells (Figures 8A, 8B, 8G, and 8H) and iNOS protein levels (Figures 8K-8L) in the brains of TBI mice.

[0084] We then investigated microglial activation and found a significant increase in Iba1-positive microglia in the cortex and hippocampus of TBI mice compared to sham controls (Figures 9A, 9B, 9E, and 9F).

[0085] This result was confirmed by Western blot of Iba1 in hippocampal extracts (Figures 9G-9H). Double-labeling experiments also showed colocalization of Iba1-positive microglia with iNOS (Figures 9A-9D). However, similar to the attenuation of astroglial activation, oral administration of GTB, but not vehicle, reduced the number of Iba1-positive astrocytes (Figures 9A-9F) and the level of Iba1 protein (Figures 9G-9H) in the brains of TBI mice. Taken together, these results suggest that oral GTB can reduce both astroglial and microglial activation in the hippocampus of TBI mice.

[0086] 10. Oral administration of GTB reduces lesion volume in the CCI model of TBI Since GTB treatment inhibited astroglial and microglial activation in the brains of TBI mice, we then decided to monitor whether oral GTB could reduce lesion volume 21 days after injury. To measure lesion volume, brain sections were stained with hematoxylin and eosin (H&E). Figure 10A represents H&E stained brain sections arranged serially to show the volume of the lesion cavity from different mouse groups. As expected, we found typical lesions with dilated cavities starting from the cortex and involving the lateral ventricles through the hippocampus in TBI mice compared to the absence of lesions in sham controls (Figure 10B). However, consistent with the suppression of astroglial and microglial inflammation, treatment with GTB, but not vehicle, reduced the size of the lesion cavity in TBI mice (Figures 10A-10B). This was also supported by quantitative analysis of lesion volume using Cavalieri stereological techniques, which revealed a reduction in total lesion volume in all hemispheres upon GTB treatment compared to either untreated or vehicle-treated TBI mice (Figure 10C).

[0087] 11.GTB treatment restores synaptic maturation in the brains of CCI-injured mice Recent studies have shown that TBI profoundly affects synaptic structure and function through a combination of instantaneous mechanical injury and resulting secondary injury processes (e.g., inflammation), ultimately leading to synapse loss. For example, according to Witcher et al., TBI induces chronic cortical inflammation mediated by activated microglia, ultimately resulting in synaptic dysfunction.

[0088] Therefore, we investigated whether GTB could protect synapses in TBI mice because GTB treatment reduced glial inflammation. PSD-95 is involved in synapse development and maturation. Double labeling of brain sections for NeuN and PSD-95 showed a loss of synapse maturation in the cortex and hippocampus of TBI mice, as indicated by a decrease in PSD-95 21 days after injury compared to sham control mice (Figures 11A-11B). On the other hand, no such loss of NeuN was observed in the cortex and hippocampus of TBI mice (Figures 11A-11B). Western blot analysis of hippocampal tissue also confirmed a marked decrease in PSD-95 in the hippocampus of TBI mice compared to sham mice (Figures 11E-11F). However, consistent with the suppression of astroglial and microglial inflammation, treatment with GTB, but not vehicle, upregulated the levels of PSD-95 in the brains of TBI mice (Figures 11A-11F).

[0089] In addition to PSD-95, other molecules such as NR2A and GluR1 are also involved in synapse maturation. Therefore, we also monitored the levels of NR2A and GluR1 and found a significant decrease in both NR2A (Figure 11E and 11G) and GluR1 (Figure 11E and 11H) in the hippocampus of TBI mice 21 days after injury compared to sham control mice. Similar to the upregulation and / or restoration of PSD-95, GTB treatment increased the levels of NR2A (Figure 11E and 11G) and GluR1 (Figure 11E and 11H) in the hippocampus of TBI mice. These results were specific since we did not observe such an increase in NR2A and GluR1 with vehicle treatment (Figure 11E, 11G&11H). These results suggest that oral GTB can restore synapse maturation in the hippocampus of TBI mice.

[0090] 12. Oral GTB protects cognitive function in TBI mice Many TBI survivors suffer from cognitive impairments throughout their lives. It has been reported that impaired synaptic changes are involved in the cognitive impairment of TBI. Because GTB treatment protected and / or improved synaptic development and maturation in the hippocampus and cortex of TBI mice, we investigated whether GTB could protect cognitive function in TBI mice 21 days after injury. To monitor short-term memory, the novel object recognition (NOR) test was used, whereas for spatial learning and memory, the behavior of mice was analyzed in the Barnes maze and T-maze.

[0091] As evident from the NOR task, TBI mice spent less time with the novel object compared to sham control mice (Figures 12A and 12C). On the other hand, when treated with GTB but not with vehicle, TBI mice spent significantly more time with the novel object (Figures 12A and 12C), indicating improved short-term memory with oral GTB. The Barnes maze is a hippocampal-dependent memory task that requires spatial reference memory. Untreated TBI mice showed less ease in finding the reward hole (Figure 12B), more errors (Figure 12D), and more time (latency) required (Figure 12E) compared to sham control mice. However, TBI mice treated with GTB but not with vehicle performed much better on the Barnes maze (Figure 12B), made fewer errors (Figure 12D), and took less time to find the target hole (Figure 12E) compared to untreated TBI mice. Similarly, in the T-maze, untreated TBI mice showed fewer positive turns (FIG. 12F) and more negative turns (FIG. 12G) than sham control mice. Consistent with the NOR task and Barnes maze, oral administration of GTB, but not vehicle, significantly enhanced hippocampus-dependent memory performance in TBI mice, as indicated by a higher number of positive turns (FIG. 12F) and a lower number of negative turns (FIG. 12G) than untreated TBI mice.

[0092] 13.GTB treatment improves motor function in TBI mice 7 days after CCI injury The primary therapeutic goal of TBI research is to maintain or restore behavioral function. Because GTB treatment protected cognitive function in TBI mice, we next investigated whether GTB also protects overall motor activity. To record general motor behavior, we used a video camera 6 (Basler Gen I Cam-Basler acA 1300-60) connected to a Noldus computer system that remained mounted facing downwards on the top of the open field activity arena. Figure 13A shows a heat map summarizing the overall movement of the mouse in the open field activity arena 7 days after CCI injury.

[0093] As expected, TBI mice showed decreased open field activity compared to sham controls in terms of heatmap (Figure 13A), distance traveled (Figure 13B), speed (Figure 13C), center frequency (Figure 13D), and rearing (Figure 13E) at day 7 after CCI injury. However, treatment of TBI mice with GTB, but not vehicle, led to a significant increase in open field behavior (Figures 13A-13E).

[0094] The motor coordination and balance activity of mice was then investigated using the roto-rod test. Similar to the open field activity, TBI mice showed a significant decrease in the latency to fall 7 days after CCI injury compared to sham controls (Figure 13F). On the other hand, oral administration of GTB, but not vehicle, improved the roto-rod performance as seen by the increased latency (Figure 13F).

[0095] Depression is a prominent symptom of TBI, especially during the early stages of brain injury, and can be monitored in mice by tail suspension test. Therefore, this test was performed to monitor the effect of GTB treatment on depression-like behavior in TBI mice. As evident from Figure 13G, TBI mice on day 7 after CCI injury showed significantly higher immobility time than sham controls, indicating more depressive behavior in TBI mice than sham mice. However, TBI mice treated with GTB showed significantly less immobility time during the tail suspension test than either untreated or vehicle-treated TBI mice (Figure 13G), suggesting inhibition of depressive behavior by GTB.

[0096] TBI is known to damage the connections between the brain and muscles, thereby impairing locomotor activity. Therefore, we used beam walking to monitor locomotor behavior and observed poorer locomotor activity in TBI mice compared to sham controls (Figure 13H-13J). TBI mice used more steps (Figure 13H), took more time (Figure 13I), and slipped more (Figure 13J) while traversing the beam than sham control mice. However, oral administration of GTB, but not vehicle, improved beam walking in TBI mice (Figure 13H-13J). To further confirm the results, we also used a grid runway, which allows scientists the opportunity to analyze and compare locomotor activity.

[0097] Similar to what was found with the beam walk, TBI mice also performed poorly on the grid runway compared to sham controls in terms of steps (Figure 13K), time taken (Figure 13L), and misplacements (Figure 13M). Again, GTB treatment improved the performance of TBI mice on the grid runway (Figures 13K-13M). Taken together, these results indicate an improvement in the motor performance of TBI mice upon GTB treatment at 7 days after CCI injury.

[0098] On the other hand, many locomotor parameters spontaneously improved on day 21 after CCI injury, and no significant changes were observed after GTB treatment (Figures 14A-14M). For example, no significant changes were observed in all parameters tested for open field behavior (Figure 14A, heatmap; Figure 14B, distance traveled; Figure 14C, speed; Figure 14D, center frequency; Figure 14E, rearing), as well as in several parameters tested for beam walking (Figure 14H, steps; Figure 14I, time taken) and grid runway (Figure 14K, steps). Only in the tail suspension test was a significant impairment found in TBI mice compared to untreated TBI mice, and GTB treatment also resulted in significantly less immobility time during the tail suspension test than either untreated or vehicle-treated TBI mice on day 21 after CCI injury (Figure 14G), suggesting that GTB can inhibit depressive behavior even in the late stages of TBI.

[0099] 14. Conclusion In summary, in a preclinical model of TBI, we demonstrated that oral GTB, a flavoring ingredient, reduced glial activation, diminished lesion cavitation, and protected cognitive and motor behaviors. Our results decipher the important neuroprotective effects of GTB and suggest that GTB may be repurposed for therapeutic intervention in TBI.

[0100] 15. Sodium benzoate (NaB) stimulates maturation of oligodendrocyte precursor cells (OPCs) into oligodendrocytes Downregulation of myelin proteins and subsequent loss of myelin sheath are believed to be pathological hallmarks of neurological conditions such as multiple sclerosis and traumatic brain injury. Therefore, we wanted to explore the effect of NaB on remyelination. Oligodendrocytes are generated from OPCs as a result of the reduction of precursor markers such as NG2 and A2B5 with subsequent induction of myelinating proteins such as myelin basic protein (MBP) and proteolipid protein (PLP). Interestingly, we found that NaB stimulated the differentiation of OPCs into oligodendrocytes (Figures 15A-15F). On the other hand, NaFO, a structural analogue of NaB, did not promote maturation of OPCs into oligodendrocytes (Figures 15A-15F). This indicates the specificity of NaB's OPC maturation effect. Accordingly, NaB treatment increased the levels of PLP and myelin oligodendrocyte glycoprotein (MOG) in OPCs (Figures 15G-15H). These results were supported by mRNA analysis of MBP, PLP, MOG, and CNPase (Figure S6I). To understand the functional significance of this finding, we investigated the effect of NaB on myelination of synthetic fibers and found stimulation of myelination by NaB but not NaFO (Figures S6J-L).

[0101] 16. Effect of NaB on remyelination in the corpus callosum in a cuprizone-intoxicated mouse model of demyelination We then investigated the effect of NaB on in vivo remyelination in the corpus callosum of mouse brain. The corpus callosum is the region that is primarily affected in various inflammatory demyelinating diseases, including MS. As expected, we found a decrease in the myelin protein PLP and an increase in the OPC marker A2B5 in the corpus callosum of cuprizone-intoxicated mice compared to control mice (Figures 16A-16C). However, NaB treatment increased the levels of PLP and decreased the levels of A2B5 (Figures 16A-16C), suggesting that NaB can promote remyelination in the corpus callosum of cuprizone-intoxicated mice. As MBP is a marker of myelin integrity, we stained corpus callosum sections with MBP and found a loss of MBP in cuprizone-intoxicated mice that increased after NaB treatment (Figures 17A and 17C). Similar results were found in the case of PLP, another stability marker of myelin fibers (Figures 17B and 17D). These results were confirmed by LFB staining (FIG. 17E) and ultrastructural details by electron microscopy (FIGS. 17F-17H).

[0102] 17. Cinnamein: Anti-inflammatory Chronic inflammation caused by macrophages, microglia, and astrocytes plays an important role in the pathogenesis of several autoimmune, inflammatory, and neurodegenerative disorders. Upon activation, macrophages, microglia, and astrocytes produce proinflammatory cytokines (tumor necrosis factor alpha or TNFα, interleukin 1β or IL-1β, interleukin-6 or IL-6, etc.) and nitric oxide (NO), which ultimately contribute to autoimmune, inflammatory injury, and neurodegenerative disorders such as rheumatoid arthritis, multiple sclerosis, Alzheimer's disease, Parkinson's disease, Huntington's disease, traumatic brain injury, etc. Therefore, the identification of non-toxic anti-inflammatory drugs may be beneficial for these autoimmune, inflammatory injury, and neurodegenerative disorders. Cinnamein is an ester derivative of cinnamic acid and benzyl alcohol, and is used as a flavoring agent and for its antifungal and antibacterial properties. Here, we demonstrate the anti-inflammatory properties of cinamein in RAW 264.7 macrophages and primary mouse microglia and astrocytes. Stimulation of RAW 264.7 macrophages with lipopolysaccharide (LPS) and interferon gamma (IFNγ) resulted in a pronounced production of NO (Figures 18A-18C). However, cinamein pretreatment for 6 h significantly inhibited LPS- and IFNγ-induced production of NO in RAW 264.7 macrophages (Figures 18A-18C). Accordingly, LPS and the viral double-stranded RNA mimetic polyinosinic:polycytidylic acid (polyIC) stimulated the production of TNFα (Figures 19A-19B), IL-1β (Figures 20A-20B), and IL-6 (Figures 21A-21B) in primary mouse microglia, which was strongly inhibited by cinamein pretreatment. Similarly, cinamein also inhibited Poly IC-induced production of TNFα and IL-6 in primary mouse astrocytes (Figures 22A-22B). These results suggest that cinamein can be used to control inflammation in a variety of autoimmune, inflammatory, and neurodegenerative disorders.

[0103] The features and advantages of the present disclosure are apparent from the detailed description, and the claims are intended to cover all such features and advantages. Many variations will occur to those skilled in the art, and any variations equivalent to those described in the present disclosure are within the scope of the present disclosure. Those skilled in the art will appreciate that the concepts on which the present disclosure is based may be used as a basis for designing other methods and systems for carrying out some of the purposes of the present disclosure. As a result, the claims should not be considered as limited by the description or examples.

Claims

1. 1. A pharmaceutical composition comprising a benzoate or a prodrug thereof for use in a method of slowing the progression of or reducing the severity of symptoms associated with nervous system injury in a subject in need thereof, the method comprising administering to the subject an effective amount of benzoate or a prodrug thereof, thereby slowing the progression of or reducing the severity of symptoms associated with nervous system injury.

2. 10. The pharmaceutical composition of claim 1, wherein the benzoate, if present, is sodium benzoate, potassium benzoate, calcium benzoate, 2-aminobenzoate, 3-aminobenzoate, 4-aminobenzoate, or any combination thereof.

3. 10. The pharmaceutical composition of claim 1, wherein the prodrug of benzoate, if present, is benzyl cinnamate, glyceryl tribenzoate, cinnamic acid, benzyl acetate, benzyl alcohol, benzoic acid, quinic acid, phenylalanine, tyrosine, or any combination thereof.

4. 10. The pharmaceutical composition of claim 1, wherein the subject has not been diagnosed with a urea cycle disorder, glycine encephalopathy, multiple sclerosis, Parkinson's disease, Alzheimer's disease, Huntington's disease, or autism spectrum disorder.

5. 10. The pharmaceutical composition of claim 1, wherein the subject has been diagnosed with nervous system damage prior to the administering step.

6. 2. The pharmaceutical composition of claim 1, wherein the nervous system injury is a central nervous system (CNS) injury or a peripheral nerve injury.

7. 2. The pharmaceutical composition of claim 1, wherein the nervous system injury is a spinal cord injury (SCI), a spinal cord contusion, or a nerve crush injury.

8. 8. The pharmaceutical composition of claim 7, wherein the effective amount of benzoate or a prodrug thereof is administered within 24 hours after SCI, spinal cord contusion, or nerve crush injury.

9. 2. The pharmaceutical composition of claim 1, wherein the nervous system injury is traumatic brain injury (TBI).

10. The pharmaceutical composition of claim 1, wherein the nervous system injury is a demyelinating disorder.

11. 11. The pharmaceutical composition of claim 10, wherein the demyelinating disorder is optic neuritis, adrenoleukodystrophy X, Krabbe disease, progressive multifocal leukoencephalopathy, adrenomyeloneuropathy, acute disseminated encephalomyelitis, acute hemorrhagic leukoencephalitis, multiple sclerosis, Barro's disease (concentric sclerosis), Charcot-Marie-Tooth disease, Guillain-Barré syndrome, HTLV-I associated myelopathy, neuromyelitis optica (Devic's disease), Schilder's disease, transverse myelitis, or a combination thereof.

12. 10. The pharmaceutical composition of claim 9, wherein the effective amount of benzoate or a prodrug thereof is administered within 24 hours after traumatic brain injury.

13. 10. The pharmaceutical composition of claim 1, wherein administering an effective amount of benzoate or a prodrug thereof results in a reduction of glial inflammation, an improvement in motor function or coordination, or an improvement in learning or memory dysfunction.

14. 10. The pharmaceutical composition of claim 1, wherein administering an effective amount of benzoate or a prodrug thereof prevents or reduces the severity of symptoms associated with depression.

15. The pharmaceutical composition of claim 1, further comprising a pharmaceutically acceptable excipient and comprising greater than 0.1% by weight of the composition of benzoate or a prodrug thereof.

16. 10. The pharmaceutical composition of claim 1, wherein the benzoate or prodrug thereof is administered orally.

17. 1. A pharmaceutical composition comprising sodium benzoate for use in a method of slowing the progression of or reducing the severity of symptoms associated with nervous system injury in a subject in need thereof, the method comprising administering to the subject an effective amount of sodium benzoate, thereby slowing the progression of or reducing the severity of symptoms associated with nervous system injury.

18. 18. The pharmaceutical composition of claim 17, wherein the subject has not been diagnosed with a urea cycle disorder, glycine encephalopathy, multiple sclerosis, Parkinson's disease, Alzheimer's disease, Huntington's disease, or an autism spectrum disorder.

19. 18. The pharmaceutical composition of claim 17, wherein the subject has been diagnosed with nervous system damage prior to the administering step.

20. 18. The pharmaceutical composition of claim 17, wherein the nervous system injury is a central nervous system (CNS) injury or a peripheral nerve injury.

21. 18. The pharmaceutical composition of claim 17, wherein the nervous system injury is a spinal cord injury (SCI), a spinal cord contusion, or a nerve crush injury.

22. 22. The pharmaceutical composition of claim 21, wherein the effective amount of sodium benzoate is administered within 24 hours after SCI, spinal cord contusion, or nerve crush injury.

23. 18. The pharmaceutical composition of claim 17, wherein the nervous system injury is traumatic brain injury (TBI).

24. 18. The pharmaceutical composition of claim 17, wherein the nervous system injury is a demyelinating disorder.

25. 25. The pharmaceutical composition of claim 24, wherein the demyelinating disorder is optic neuritis, adrenoleukodystrophy X, Krabbe disease, progressive multifocal leukoencephalopathy, adrenomyeloneuropathy, acute disseminated encephalomyelitis, acute hemorrhagic leukoencephalitis, multiple sclerosis, Barro's disease (concentric sclerosis), Charcot-Marie-Tooth disease, Guillain-Barré syndrome, HTLV-I associated myelopathy, neuromyelitis optica (Devic's disease), Schilder's disease, transverse myelitis, or a combination thereof.

26. 24. The pharmaceutical composition of claim 23, wherein the effective amount of sodium benzoate is administered within 24 hours after traumatic brain injury.

27. 18. The pharmaceutical composition of claim 17, wherein administering an effective amount of sodium benzoate results in a reduction of glial inflammation, an improvement in motor function or coordination, or an improvement in learning or memory dysfunction.

28. 18. The pharmaceutical composition of claim 17, wherein administering an effective amount of sodium benzoate prevents or reduces the severity of symptoms associated with depression.

29. The pharmaceutical composition of claim 17, further comprising a pharmaceutically acceptable excipient and comprising more than 0.1% by weight of the composition of sodium benzoate.

30. 18. The pharmaceutical composition of claim 17, wherein the sodium benzoate is administered orally.

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