Arches. Photo by Daniel Chia
HOPES: Huntington's Outreach Project for Education, at Stanford
Jun
29
2010

Minocycline

Update: a 2010 study of Minocycline, called DOMINO, concluded that minocycline does not warrant further study for the treatment of HD. 114 patients who had mild to moderate HD were treated with either 200 mg of minocycline or a placebo every day for 18 months. The study measured patient’s improvement in the Total Functional Capacity (TFC), a test that measures an HD patient’s ability to function in day-to-day life. The conclusion – that minocycline is not worth studying further – is unfortunate, but clears up resources for the study of other drugs.

DOMINO was a futility study, which is a small trial that is designed to show whether a drug is worth the expense of a full clinical trial. The standards for a futility study are low; the study only needs to show that there is a reasonable chance that the drug is more effective than a placebo. The scientists performing the study attempted to show that minocycline could be 25% better than placebo – a modest goal – but concluded that there is only a small chance that minocycline can reach even that threshold. Therefore, minocycline is unlikely to be considered for further trials. For more information, click here.

The study: Huntington Study Group DOMINO Investigators. A futility study of minocycline in Huntington’s disease. Mov Disord. 2010 Oct 15;25(13):2219-24.

Drug Summary: Studies have discovered that minocycline is able to inhibit the activation of cells involved in inflammation as well as decrease the production of free radicals. Because long-term inflammation as well as increased free radical production are believed to contribute to the progression of HD, minocycline treatment may aid in delaying HD progression.

Minocycline and Inflammation^

Minocycline, an antibiotic commonly used to treat acne and some forms of arthritis, has been found to delay disease progression and mortality in mice with Huntington’s disease (HD). Minocycline is able to cross the blood-brain barrier and exhibit anti-inflammatory effects. The blood-brain barrier is a group of cells that form a special, selectively permeable lining in the blood vessels of the brain. This lining serves to prevent toxic substances in the blood from entering the brain.

The anti-inflammatory effects of minocycline include inhibition of microglial activation. Microglia are cells found in the brain that are involved in the immune response. By inhibiting the activation of microglia, minocycline inhibits inflammation. (For more on microglia cells and HD, click here)

Minocycline and Free Radical Formation^

In addition, minocycline has been found to decrease free radical formation. Free radicals are very reactive molecules that are capable of inducing various biological damage. Studies have reported that free radicals play a role in the progression of HD. Minocycline decreases free radical formation by inhibiting the production of inducible nitric oxide synthetase (iNOS), an enzyme responsible for the formation of nitric oxide (NO), which acts as a free radical in the cell. Increased iNOS activity is present in activated glial cells of the HD brain. As iNOS activity increases, more nitric oxide molecules are produced, resulting in greater damage from free radicals. Figure 1 shows an illustration of how minocycline decreases free radical production.

Minocycline and Protein Aggregation^

Minocycline also inhibits the production of caspases, a family of enzymes involved in HD progression. Evidence suggests that caspases are important mediators of inflammation and apoptosis. Caspases are activated in the brains of humans with HD and certain mouse models of HD. Once activated, caspases cleave the altered huntingtin protein. Studies have shown that the cleaved huntingtin fragments easily aggregate to form aggregations called neuronal inclusions (NIs) that are toxic to the cell.

Caspases are also required for the processing of mature Interleukin-1 (IL-1), one of the cytokines involved in the inflammatory response. Cytokines are one of the “weapons” by which the immune system removes and kills foreign substances. Mature cytokines such as IL-1 then go on to initiate various pathways that further increase cellular damage. By inhibiting caspases, minocycline could potentially decrease formation of neuronal inclusions as well as inflammation in the brain.

Because inflammation, free radicals, and neuronal inclusions are all believed to contribute to the progression of HD, it is possible that minocycline may work on these various pathways and decrease or delay nerve cell death in people with HD. The success of minocycline as a treatment for HD in animal models has not yet been applied to clinical trials on humans, but pilot studies to test the safety/tolerability and efficacy of minocycline in patients with early HD are currently underway.

Research on Minocycline^

Chen, et al. (2000) investigated the overall effects of minocycline on HD progression. They hypothesized that minocycline should act on the various disease mechanisms associated with HD and cause a delay or improvement in the condition of treated mice.

To assess the efficacy of minocycline, the researchers looked at changes in motor performance, survival rates, and amount of huntingtin fragments and neuronal inclusions.

The investigators gave injections of minocycline to 6-week old mice that expressed the symptoms of HD. At 6 weeks of age, the mice were in the early stages of HD and showed some of the pathological characteristics associated with HD, such as NI formation and declined motor performance. The researchers found that minocycline significantly delayed the decline in motor performance. Furthermore, the treated mice lived 14% longer than untreated mice. This extended period of survival is roughly equivalent to one to five years in people with HD.

As a means of comparison with minocycline, the researchers also injected tetracycline to another group of mice. Tetracycline is a drug similar in effects to HD but is not known to cross the blood-brain barrier. Mice treated with tetracycline showed no improvements in performance or survival.

Minocycline-treated mice were also found to have significantly lower levels of the huntingtin fragments. However, minocycline did not inhibit formation of neuronal inclusions (NIs) despite the fact that lower levels of huntingtin fragments are present in the brains of minocycline-treated mice. (For more on neuronal inclusions, click here)

These results indicate that minocycline-mediated neuroprotection is not related to the effect that NIs have on the disease. The researchers believe that NI formation is dependent on caspase activation only during the initial stages of huntingtin aggregation. Specifically, the cleavage of the full-length huntingtin may be the caspase-dependent step. Once cleavage occurs, aggregation of the fragments and consequent formation of NIs follows.

Once NIs begin to form, their growth is no longer dependent on caspase activity since the formation of aggregates no longer require caspase cleavage. In short, caspases produce the fragments needed to form NIs but are not needed in the aggregation process of these fragments. NIs have been detected in HD mice as early as three weeks of age. The minocycline administered in this study probably did not inhibit NI formation because it was administered when the mice were already six weeks old. At that time, the early caspase-dependent step was likely to have passed and aggregates were already able to form without the need for caspase-activity.

Minocycline treatment also resulted in a 72% inhibition of iNOS activity in brains of HD mice when compared to untreated HD mice. These results indicate that neuroprotection from minocycline treatment results in part from inhibition of iNOS activity, which leads to decreased free-radical damage.

Researchers are not exactly sure how caspases are activated or inhibited. They do know that caspases are not always present in the cells and are produced only during specific times such as inflammation or apoptosis. Beginning in the early stages of HD, the altered huntingtin induces caspase production and activation, resulting in mature IL-1 production and huntingtin cleavage. As activated caspases play a role in HD progression, an effective therapeutic intervention would require inhibition of these various caspases.

Minocycline has been shown to inhibit caspase production, though the mechanism by which inhibition occurs is still not known. Minocycline also inhibits the formation of nitric oxide, making it an important neuroprotective compound. Few side effects have been reported by people treated with minocycline, as it is a fairly safe and common antibiotic used to treat diseases such as acne and arthritis. With the results of this study and its low toxicity, minocycline represents a new potential therapeutic agent for HD treatment.

Tikka, et al. (2001) studied the effects of minocycline on microglial proliferation due to NMDA receptor activation. (For more on NMDA receptors, click here.) The researchers hypothesized that minocycline treatment to cells exposed to excitatory molecules such as NMDA can protect nerve cells from death.

NMDA receptors are activated by various excitatory molecules such as NMDA and glutamate. Activation of these receptors leads to entry of calcium ions (Ca2+) into the cell. Ca2+ entry then activates various calcium-dependent proteins and molecules that can initiate activities promoting cell death. The researchers believe that NMDA receptor activation also leads to microglial proliferation and activation of the inflammatory response. In the rat brain, the microglia cells are known to have various NMDA receptors on their surfaces, explaining why increased glutamate levels trigger microglial proliferation and activation. Increased glutamate levels or NMDA activation (as known to happen in HD cells) can therefore induce microglial proliferation and activation.

Minocyline, a compound known to decrease the activation of microglia, was used to study whether it will have any beneficial effects on cells exposed to NMDA. To test the efficacy of minocycline, the researchers looked at the amount of activated microglia and the survival rates of nerve cells.

The researchers exposed a group of nerve cells to NMDA. They then added minocycline to one group of nerve cells while left another group untreated. Following NMDA administration, the researchers saw an increase in microglial proliferation, followed by an increase in nerve cell loss. These changes were also associated with increased release of cytokines and nitric oxide free radicals. Minocycline administration was found to reduce microglial proliferation and nerve cell loss.

The researchers also discovered that increased microglial activation enhances the neurotoxicity of NMDA. They believe that the enhanced toxicity arose due to the increased release of cytokines and free radicals from the activated microglia. Increased cytokine production has been known to result in delayed removal of glutamate molecules, enhancing NMDA receptor activation.

One way by which the researchers believe that minocycline was able to reduce nerve cell loss was by its ability to inhibit caspases. Because caspases are needed for the maturation of certain cytokines, inhibiting the caspases could have decreased damage due to the cytokines. Decreased production of the mature cytokines would have enabled the normal removal of glutamate and excitatory molecules, thereby decreasing NMDA receptor activation. However, the researchers are still uncertain as to how minocycline was able to reduce the proliferation of microglia.

In conclusion, the study showed that minocycline treatment results in the inhibition of microglial activation and decreased release of cytokines and certain free radicals. Minocycline may therefore be beneficial to people with diseases such as HD that involve toxicity due to excitatory molecules, although more research is needed.

For further reading^

  1. Chen, et al. “Minocycline Inhibits Caspase-1 and Caspase-3 Expression and Delays Mortality in a Transgenic Mouse Model of Huntington Disease.” Nature Medicine. 2000; 6:797-801.
    This study reported that minocycline was able to reduce huntingtin fragments and nitric oxide formation, but had no effect on neuronal inclusion formation.
  2. Tikka, et al. “Minocycline Provides Neuroprotection Against N-Methyl-D-aspartate Neurotoxicity by Inhibiting Microglia.” The Journal of Immunology. 2001; 166: 7527-7533.
    This study reported that minocycline treatment is associated with reduced inflammation and nerve cell loss due to NMDA activation.

-P. Chang, 5/6/03, updated by M. Hedlin on 6/29/11