Potential Use of Calpain Inhibitors as Brain Injury Therapy

Excerpt

Existing therapies for traumatic brain injuries (TBI) only relieve the symptoms of patients but do not treat the underlying causes that lead to debilitating effects of the trauma. One of the well understood molecular mechanism of TBI is the hyperactivation of the calpain enzyme. The need for drugs to inhibit the effect of the enzyme is viewed as one of the neurotherapies for traumatic brain injury. In this chapter, we summarize one of the strategies in finding small molecules to slow down or even eliminate the devastating effect of calpain hyperactivation.

The brain is a well-protected organ. Because of the very fragile nature of the brain, the organ is encased in a series of bone, membrane, and fluid-based protection. But despite all the privileges the brain is bestowed with, the organ is still very vulnerable to damage. A car accident, a football tackle, and an explosive’s shock wave from a blast are all blows to the head that can set off a chain reaction, rendering the victim at least incapacitated or at worst, dead. At the molecular level, it is now well-documented that part of the damage is from the hyperactivation of calpain.

Calpain is a tightly regulated Ca²⁺-dependent proteolytic enzyme that plays a critical role in important signaling pathways, including synaptic function and memory formation. Calpain exerts its biological function through limited proteolysis of its substrates. However, in some pathological conditions, such as after a TBI, calpain is dysregulated and participates in cell injury and death.

In TBI, neuronal calpains are hyperactivated within minutes after the initial trauma event, causing early damage to specific areas of the brain (Huh et al., 2007; Kampfl et al., 1996; Liu et al., 2006; Pike et al., 1998, 2001; Posmantur et al., 1997; Thompson et al., 2006; Upadhya et al., 2003; Warren et al., 2005; Zhang et al., 2007; Zhao et al., 1998a, 1998b, 2000). In multiple models of TBI, sustained calpain activation is associated with neuronal and axonal cell death. Calpain is not just linked to TBI but also to a handful of neurogenerative diseases. These neurogenerative pathologies include cerebral ischemia (Blomgren et al., 1999; Garcia-Bonilla et al., 2006; Gutierrez et al., 2007; Hong et al., 1994; Johnson et al., 1997; Kambe et al., 2005; Liebetrau et al., 1999; Minger et al., 1998; Neumar et al., 1996; Tsubokawa et al., 2006), Alzheimer’s disease (DiRosa et al., 2002; Garg et al., 2011; Getz, 2012; Grynspan et al., 1997; Iwamoto et al., 1991; Medeiros et al., 2012; Tsuji et al., 1998), multiple sclerosis (Das et al., 2008; Shields et al., 1999), Huntington’s disease (Bizat et al., 2003; Gafni and Ellerby, 2002; Gladding et al., 2012), prion-related encephalopathy (Gray et al., 2006; Hachiya et al., 2011; Wang et al., 2005; Yadavalli et al., 2004), and Parkinson’s disease (Alvira et al., 2008; Mouatt-Prigent et al., 1996; Ray et al., 2000; Samantaray et al., 2008).

Enzyme inhibition is a well-known strategy for therapy development, and based on our understanding of calpain in the pathology of brain injury, this enzyme is an attractive therapeutic target. Calpain inhibitors probably came out at the same time as the discovery of the said enzyme 40 years ago. However, these inhibitors were general protease inhibitors, lacking selectivity and therefore inhibiting other proteases such as the proteasome and cathepsins. Interest in making selective calpain inhibitors sprung from the need to pinpoint the specific role(s) of calpain in the body. Subsequent years of research in calpain inhibitors led to a variety of compounds, most of which are small molecules (average MW ~500 g/mole).

As drug candidates, small-molecule compounds have some advantages over peptides or proteins, such as the ability to penetrate cells and resistance to proteolytic degradation. However, peptide and protein drug candidates exhibit higher selectivity toward their targets. A desirable treatment strategy in a clinical setting is through oral administration with a convenient dosing schedule, probably twice a day frequency of ingestion. These desirable pharmacological properties of a drug that can be taken orally and dosed once or twice daily are all met by small molecule compounds. In addition, small molecules are simpler: only a small portion of the enzyme, the active site is targeted to render the enzyme unable to perform its function. Compared with the whole enzyme, the active site is relatively small in comparison to the total volume of the macromolecule. This is akin to poking a small hole in the gas tank of a car so that the whole automobile cannot function.

The active site is precisely arranged in a three-dimensional structure to facilitate the catalysis of substrates. This is generally where the action occurs, and rendering the active site unable to perform its duty (or partially incapacitated) is one of the goals of small molecule inhibitors.

Designing small molecules as inhibitors is a strategy that is common in the pharmaceutical industry. In the market today, small-molecule enzyme inhibitors comprise half of all drugs. Pharmaceutical companies will still continue to follow in that direction, developing small molecules to inhibit dysregulated enzymes. And there are relatively a lot of success stories in drug development that targets enzymes. An example of this is the angiotensin-converting enzyme (ACE) inhibitor for treating hypertension. ACE converts angiotensin I into angiotensin II, resulting in blood vessel constriction and eventually increasing blood pressure. Blocking ACE lowers blood pressure. Pfizer’s Viagra, a drug that blocks the enzyme cyclic guanosine monophosphate–specific phosphodiesterase type 5 to combat erectile dysfunction, is another blockbuster success. There has been a focus on designing new inhibitors of proteases that have been identified as disease-causing proteolytic enzymes. In addition to the successfully inhibited enzymes mentioned earlier, several inhibitors of a number of proteases are believed to progress from purely research to clinical trials. Bayer has developed a cathepsin K inhibitor (AAE581) to treat osteoporosis (Falgueyret et al., 2005). We saw the development of a serine aminopeptidase inhibitor, vildagliptin, as a therapy for type 2 diabetes (Deacon and Holst, 2006), and another ACE inhibitor (Aliskiren) that is an alternative to the current ACE inhibitor drugs (Stanton, 2003).

In this chapter, we will address the issues and challenges in drug discovery, specifically for the viability of small molecule inhibitors to regulate the destructive effects of calpain hyperactivation during or after TBI. Broadly, the enzyme (2) calpain will be discussed, followed by (3) calpain inhibitors, (4) challenges in realizing a therapy for brain injury, and then THE (5) conclusion. We hope that this chapter will provide students and scientists in brain injury research an overview of the potential of small-molecule calpain inhibitors in TBI drug development.