Alzheimer's Disease

AD is a slowly degenerative brain disease that impairs memory, attention and judgment. Most people with AD are age 65 or older, and the risk of acquiring the disease increases with age. AD is the most common form of dementia in older people. Estimates according to the National Institute on Aging say that approximately 3 percent of people ages 65-84 have AD and nearly half of those over the age of 85 could have the disorder. Four and one half million people in the United States have this progressive, degenerative brain disorder. Although the risk of getting AD increases with age, it is not a normal part of aging.

Groups of neurons are located throughout the brain. In AD, the areas of the brain that are the most affected are the hippocampus and the cerebral cortex, the outer layer of the brain. The areas of the cerebral cortex for cognitive functions such as language are most affected in AD. The hippocampus, which is located deep inside the brain, is believed to play an important role in memory. Death of neurons in these important parts of the brain has a severe impact on memory, cognition, and behavior in the AD patient. Calcium has been overwhelmingly shown as a large contributor to AD and related dementias. Our goal is to reduce the neurotoxic effects of calcium by removing the calcium. Disturbances in calcium homeostasis have been observed to be associated with AD and other neurodegenerative diseases. Increased total calcium levels and reductions of calcium-binding proteins (calbindin-28k and calmodulin) have been found in AD brains.

In Alzheimer's disease (AD), abnormal accumulations of Aß are present in the brain and degenerating neurons exhibit cytoskeletal aberrations (neurofibrillary tangles). Roles for Aß in the neuronal degeneration of AD have been suggested based on recent data obtained in rodent studies demonstrating neurotoxic actions of Aß. Aß protein is thought to underlie the neurodegeneration associated with AD by inducing Ca(2+)-dependent apoptosis. Beta amyloid toxicity in culture is accompanied by multiple events culminating in apoptosis. One of the postulated mechanisms of beta-amyloid toxicity seems to involve a Ca2+ dysregulation accompanied with enhanced vulnerability to excitotoxic stimuli.

Parkinson's Disease (PD)

PD is a disorder known for its characteristic tremors or muscle spasms, which can be mild or severe enough to hamper mobility. Some Parkinson's patients display the same kind of confusion and memory impairment as patients with AD. The affected area in the brain with PD is the substantia nigra. This part of the brain releases a neurotransmitter called dopamine. A rise in intracellular calcium concentrations represents one of the final events leading to nerve cell death. Calcium-binding proteins have been suggested as having a neuroprotective role in dopaminergic cell groups. PD is a disorder, in which neurons of various neuronal systems degenerate. Furthermore, in such degenerating neurons, the cytoskeleton seems to be affected. In this respect, PD resembles AD. It has been shown that elevated levels of intracellular calcium can disrupt the cytoskeleton and that the stimulation of glutamate (NMDA) receptors can cause high intracellular concentrations of calcium. It has therefore been suggested, that the stimulation of glutamate receptors plays a role in the slow degeneration in AD and PD. In the case of the degeneration of the dopaminergic nigrostriatal system in PD, neurons that contain CaBPs appear to be less vulnerable than the neurons that lack it, suggesting that CaBPs might protect these neurons from degeneration by preventing that cytosolic calcium concentrations increase excessively.

Amyotrophic Lateral Sclerosis (ALS)

Introduction

Quincy Bioscience is developing therapeutics based on the calcium-binding protein apoaequorin to treat ALS and other neurodegenerative conditions/diseases. We feel that given the neuronal susceptibility to calcium-mediated toxicity that the calcium buffering properties of apoaequorin will offer an amount of neuroprotection to the areas of the brain that are known to be lacking endogenous calcium-binding proteins. It is hoped that through the control of excessive calcium that other neurotoxic events are also deterred.

Calcium and ALS

In ALS the neurons that control muscular movements are subject to degeneration. Early signs include weakness of the hands, along with muscle atrophy. The weakness and atrophy slowly creep up the forearms to the shoulders and the lower limbs can become weak and spastic. Rapid-twitching (fibrillation) is almost always present and death is a result of the eventual atrophy of respiratory muscles. It has been hypothesized (Elliot, 1995 [1] that failure of specific glutamate transports and calcium-binding proteins may account for the selective vulnerability of the corticomotoneuronal system. It is also now widely accepted that the distribution of calcium-binding proteins (parvalbumin and calbindin-D28k) play a neuroprotective role in motoneuron diseases that are a result of disturbance in the neuronal calcium homeostasis (Ince et al., 1993 [2]). The lack of calcium buffering proteins parvalbumin and calbindin D-28k and the low expression of the GluR2 AMPA receptor subunit may render human motoneurons particularly vulnerable to calcium toxicity following glutamate receptor activation (Shaw et al., 2000 [3]). Experimental studies have suggested that increased calcium and inappropriate calcium handling by motoneurons might have a significant role in motoneuron degeneration (Alexianu et al., 2000 [4]). Parvalbumin has been shown in ALS-resistant motoneurons (Elliot and Snider, 1995 [5]; Alexianu et al., 1994 [6]) along with calbindin -D28k (Alexianu, 1994 [7]). What this implies is that in areas of the brain where calcium-binding protein are found, the neurons are protected from degeneration in ALS. ALS may be caused by excitotoxic neuronal cell death (Appel, 1993 [8]) and Ca(2+) poisoning (Brown, 1994 [9]). In ALS it has been demonstrated that Ca(2+) is increased in the spinal cord of patients and that this intracellular Ca(2+) is the key mediator in the process of cell degeneration (Appel, 1993 [10]). Apoaequorin could provide a neuroprotective role in ALS by buffering calcium ions that cannot be handled correctly due to a deficiency in endogenous calcium-binding proteins. There is also a heightened vulnerability of neurons that normally express high somatodendritic levels of neurofilament (e.g., entorhinal and association cortices in AD, the spinal cord in a mouse model of ALS, and the retina in a primate model of glaucoma), as well as the reduced vulnerability of neurons that express calcium-binding proteins (e.g., neocortex of AD patients, the spinal cord and brainstem of ALS patients, and the spinal cord of a mouse model of ALS) (Morrison, et al. 1998 [11]). Motorneurons are particularly vulnerable both in human forms of ALS and corresponding animal models of the disease. With respect to motoneuron degeneration, measurements suggest that the exceptional stability of oculomotoneurons partially results from a specialized calcium homeostasis based on high buffering capacities (Vanselow and Keller, 2000 [12]). The lack of SOD-1 may be associated with vulnerability to insult by depletion of non-mitochondrial calcium stores selectively in motoneurons lacking parvalbumin and/or calbindin D-28k (Siklos et al., 2000 [13]). The calcium-binding protein parvalbumin is found in neurons that are resistant to degeneration in amyotrophic lateral sclerosis (ALS) (Elliot and Snider, 1995 [14]; Alexianu et al., 1994 [15]). It is also now widely accepted that the distribution of parvalbumin plays a neuroprotective role in motoneuron diseases that are a result of disturbance in neuronal calcium homeostasis (Ince et al., 1993 [16]). The lack of calcium buffering proteins parvalbumin and calbindin D-28k and the low expression of the GluR2 AMPA receptor subunit may render human motoneurons particularly vulnerable to calcium toxicity following glutamate receptor activation (Shaw et al., 2000 [17]).

Proposed Experiments

Quincy Bioscience's studies are designed to begin evaluating the hypothesis that the naturally occurring calcium binding protein apoaequorin can provide an important boost to the buffering capacity of older neurons in the brain and that such effects will improve neuronal and overall brain function in aged animals, including humans. In order to test the potential benefits of apoaequorin, it is necessary to begin by investigating the feasibility of introducing apoaequorin into neurons in a series of in vitro and in vivo studies. After evaluating the effects of apoaequorin on neuron survival, the long-term effects of apoaequorin treatment will be studied to demonstrate (a) how long it remains in neurons and (b) whether there are any negative side effects with long-term exposure to apoaequorin.

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Bibliography
[1] Elliot, 1995 [2] Ince et al., 1993 [3] Shaw et al., 2000 [4] Alexianu et al., 2000 [5] Elliot and Snider, 1995 [6] Alexianu et al., 1994 [7] Alexianu, 1994 [8] Appel, 1993 [9] Brown, 1994	[10] Appel, 1993 [11] Morrison, et al. 1998 [12] Vanselow and Keller, 2000 [13] Siklos et al., 2000 [14] Elliot and Snider, 1995 [15] Alexianu et al., 1994 [16] Ince et al., 1993 [17] Shaw et al., 2000

Huntington's Disease

Huntington's disease (HD) is dependent upon the correct functioning of the basal ganglia. The degeneration of neurons in the basal ganglia is thought to be due to excitotoxic activity related to an increase in intracellular calcium. In HD there is significant loss of calbindin-containing neurons in the basal ganglia.

Pick's Disease

Similar to other neurodegenerative disorders, Pick's disease is a result of neural toxicity due to excess calcium and calcium-mediated events.