Oxidative Damage and Valproic Acid
As evidenced by the Cartier et al. (2009) study, the severity of CALD provides a reason to attempt treatments that are experimental and pose risks. Oxidative stress likely plays a large role in the onset of CALD, among many other neurodegenerative diseases such as Alzheimer’s Disease and Parkinson’s Disease. Thus, understanding and treating oxidative damage in CALD is a route that needs to be explored because it can provide a great deal of research for X-ALD that has applications for a number of other diseases.
The mechanism by which VLCFA accumulation causes oxidative stress is still poorly understood. Since ROS are formed mainly by the mitochondria, there has been some research that addresses this issue. For example, a recent study has shown that VLCFA interferes with mitochondrial DNA and inhibits oxidative phosphorylation (López-Erauskin et al., 2013). The way in which VLCFA damages the DNA, however, remains just speculation. Experiments aimed at elucidating this mechanism could include an altered version of the novel assay made by Wiesinger et al. (2013). They were able to isolate peroxisomes from the cytosol so it should be possible to isolate mitochondria. If VLCFA concentrations found in X-ALD patients are enough to induce ROS from the mitochondria, this will provide insight into whether VLCFA accumulation is directly responsible for mitochondrial dysfunction or if there are intermediate factors that lead to oxidative stress.
HDAC Inhibition as a Therapeutic Target for CALD
Since ABCD2 has been shown to mitigate oxidative damage, upregulation of ABCD2 expression could prove to be a valuable therapeutic approach. This upregulation has been accomplished by the histone deacetylase (HDAC) inhibition activity of valproic acid (Fourcade et al., 2010). Further research into the viability of valproic acid (Figure 1) as a therapeutic agent needs to be explored in a clinical trial. Since it is already an FDA-approved drug, this could provide a more rapid response to treating CALD than developing a new drug.
Since valproic acid is an indiscriminate HDAC inhibitor, it may be less effective than other drugs, which selectively inhibit certain classes of HDAC. Vorinostat is a small molecule that has been used in the treatment of cancer (Figure 2). It has been shown to inhibit both VLCFA accumulation and proinflammatory cytokines in X-ALD fibroblasts and mouse model X-ALD (Singh et al., 2011). Whether this just upregulates ABCD2 expression as valproic acid does, however, is not known. It would be very useful to study the effects of vorinostat on gene regulation as well as if it does have effects in mitigating VLCFA-induced oxidative stress. Using gene expression profiling, it would be possible to understand what other factors are being influenced by valproic acid.
Adenoviral Gene Therapy
The effectiveness of the Cartier et al. (2009) gene therapy methodology was sufficient to reduce the symptoms of CALD to a level similar to those from donor marrow transplants. There are inherit problems with lentiviral vectors, however, as they incorporate into the host cell’s genome, which can have adverse effects if the integration occurs in the middle of an important gene. Another common viral vector system for gene therapy is adenoviruses. Genes from adenoviruses are found in the nucleus of the infected cells, and they are able to be expressed without incorporation into the host’s genome. There is a new type of adenoviruses that are called helper-dependent adenovirus (hdAd) vectors that have shown a lot of promise in gene therapy (Wong et al., 2013).
The hdAd vectors are able to infect cells without infecting them with any sort of viral protein coding sequences (Wong et al., 2013). This most likely helps circumvent the cell’s anti-viral response. The genes are then expressed without the risk of disrupting normal genetic function. The hdAd vectors have been shown to produce high levels of expression of the transduced genes . This means that microglia would be able to express higher quantities of the ABCD1 gene so that a lower percentage of engraftment would be necessary. This would allow for less severe ablation of the CALD patient’s bone marrow cells, which could minimize the health risks associated with bone marrow transplant.
X-ALD is characterized by VLCFA accumulation in cells that is a direct consequence of mutations in ABCD1. This increase in VLCFA has several proposed effects (Figure 3). First, it is incorporated into cholesterol esters, and this is likely to destabilize the membrane of myelin. Microglia, then, try to phagocytize the VLCFA that are released, but they are unable to degrade them. This causes an inflammatory response that is responsible for the inflammatory demyelination found in CALD. Second, since myelin has a greater concentration of VLCFA than most cells, the oligodendrocytes that create the myelin sheath must also have more VLCFA than normal cells. This causes them to be very susceptible to loss of ABCD1 function, and they undergo apoptosis. The myelin membrane degradation caused by VLCFA accumulation, thus, cannot be repaired. This is compounded by the fact that any residual repair that oligodendrocytes could manage is not happening because microglia apoptosis precludes its ability to activate oligodendrocytes. Third, VLCFA induce oxidative stress on the cell that can cause oxidative damage as well as destroy AMPK, which has known anti-inflammatory effects.
Focus on the Future
The most important question in X-ALD research is this: what triggers the inflammatory response in some people and not others? There are many proposed theories that seem to have solid biochemical and cell biological bases. For example, microglia have been shown to secrete proinflammatory cytokines and chemokines because of their inability to degrade VLCFA. It is not clear why this only occurs in some patients. It alsoe does not explain, however, why head trauma can induce CALD. The infiltration of lymphocytes in the latter case may be due to structural integrity issues with the blood brain barrier (Raymond et al., 2010). So, if lymphocyte infiltration is observed without trauma, it is possible that there may be other factors that increase the recruitment of inflammatory factors.
With the observation that AMPK is destroyed by the ubiqutin-proteasome system, the link between oxidative damage and inflammatory response becomes somewhat clearer. This, however, only shows that oxidative stress stops AMPK from activating its anti-inflammatory effects (Singh and Giri, 2014). The probability that oxidative stress is causing the upregulation of inflammatory factors remains likely, but not well documented.
Understanding the epigenetic and environmental factors that trigger the inflammatory response is of great importance for X-ALD treatment. While bone marrow transplant is an effective treatment for CALD, it poses a number of serious health risks so it should not be used as a preventive measure—only when it is clear that a person is presenting with the CALD phenotype. Thus, finding the factor or number of factors that cause the shift to the rapid, inflammatory demyelination that characterizes CALD must be the focus of X-ALD study.