By David N. Leff

Neuropathologists beating the bushes for the perpetrator of Alzheimer's disease (AD) have four top suspects - plaques, tangles, APOE-4 and tau:

¿ Senile neuritic plaques in the brain surround nerve cells involved in memory and cognition. But that circumstantial evidence isn't enough to convict.

¿ Neurofibrillary tangles hide out inside those neurons, but again, no smoking gun.

¿ APOE-4 (apolipoprotein E-4) acts more like a cut-out than a co-conspirator. On the AD most-wanted list it rates as a "risk factor"- a not-so-innocent bystander.

¿ And then there's the shady, slippery tau protein, (unrelated to the Greek letter), which seems to be playing the AD criminals and crime-hunters off against each other.

Cell biologist Kun Ping Lu, who is on the medical staff of Harvard-affiliated Beth Israel-Deaconess Hospital, in Boston, has much of the goods on tau.

"It's one of the proteins that binds microtubules," he told BioWorld Today. "Their cytoskeleton network plays an important role in the structure and function of a neuron." Lu explained: "Neurons have a distinct structure. They have a long axon and short dendrites. The axons exist in long structures that are maintained by the microtubules. These microtubules also play an important part in the proteins that are synthesized in the soma - the neuron's cell body. They transport nutrients and building materials all the way down to the synapse at the tip of the axon, which could be a meter away from the cell body inside the spinal cord. Some such neurons direct muscle contraction in the leg."

Tau is one of those construction proteins. It assembles tubulin, the microtubules' building blocks, into that neuronal cytoskeleton network. The gene that expresses tau protein resides on the long arm of human chromosome 17. When that gene is multiply mutated, some of the altered tau proteins become hyperphosphorylated. That is, they acquire an overabundance of phosphate. This changes the conformational shape of tau, and disrupts its job of binding and assembling tubulin.

"So tau cannot perform its function," Lu pointed out, "and tubulin can collapse. So now the neuron doesn't have this cytoskeleton network any more. At the same time, hyperphosphorylated tau will tangle itself to form a paired helical filament - that is, a neurofibrillary tangle. That tangle is a rigid structure, present in the neural cell body, and also in its dendrites and axons. And these tangled structures will block the function of the neuron, damage it and - according to one hypothesis - eventually cause its death."

Lu is senior author of a paper in today's Nature, dated June 24, 1999, titled: "The prolyl isomerase Pin1 restores the function of Alzheimer-associated phosphorylated tau protein."

He discovered Pin1 four years ago while a postdoctoral fellow at the Salk Institute for Biological Studies, in La Jolla, Calif. "The Pin1 enzyme," Lu said, "is ubiquitously expressed in all human tissues and cells we have so far examined. It's expressed by a gene on the short arm of chromosome 19, and plays a vital role in all cell division. Pin1 is an enzyme that changes the conformation of specific phosphorylated proteins. Just how specific is the fact that it only affects the changes in shape of two essential amino acids, serine and threonine, and only when those residues immediately precede proline, a nonessential amino acid."

"Proline," Lu said, "has the ability to put a kink into certain protein structures. If you put phosphate on the serine or threonine before the proline residue, now you have made two distinct conformations. One form is a kinky structure, the other, an extended one. And Pin1 can change this conformation - switch from one form to another."

Lu has confirmed this property of the enzyme in experiments that compared its behavior in healthy human brain autopsies with those of AD patients.

"We found that in a healthy brain," he said. "Pin1 is pharmacologically available, because it's in soluble form. The enzymes are in the cell nucleus where they should be and perform their function. But, in AD, Pin1, instead of being in the cell's nucleus, is present in the tangle, located in the cytoplasm. And more important, it's insoluble and unavailable, whereas available Pin1 gets depleted - apparently from working overtime trying to fix tau. By this experiment, we found that in the normal brain, 90 percent of the Pin1 protein is in the soluble fraction. But, in an AD brain, 90 percent is in the insoluble fraction."

Lu said "a lot of drug companies are now trying to develop drugs for AD by finding a compound that would block phosphate from attaching to tau. But our study offers an entirely new idea, and opens a completely novel avenue for therapy. We achieve the same function. We can alter the function of tau protein by changing its conformation with Pin1, even if the tau is hyperphosphorylated. So one strategy would be to try somehow to up-regulate our enzyme, and allow it to restore tau's function."

Patents Allowed And Pending

Lu allowed that he and his co-workers are still far away from this goal. "A lot of experiments have yet to be done," he said. "I think, first, we have to prove that we can find a way to up-regulate Pin-1. Second, after we do that, either slow down or prevent tangle formation. Third, [we can] thus prevent neuronal death." These in vivo experiments are now under way in his lab, on mouse models."

One U.S. patent allowed to the Salk Institute protects the Pin1 intellectual property, while three other pending applications (two held by Beth Israel-Deaconess Hospital) cover the enzyme's inhibitors and modulators. Lu is now in discussions "regarding drug development with several pharmaceutical and biotechnology companies," he said. "In AD, we are trying to up-regulate Pin1. But this enzyme, potentially, could be a drug target for other diseases. Because it controls cell division, the Pin1 inhibitor can potentially be used for treating cancers."