23 February, 2013

Amyloid and Memory

"Whats the last thing that goes through a bug's mind as it hits the windshield of your car?"

The answer to this old joke ("it's a**hole") may turn out only to be true if said bug has defective Orb2, a member of a family of RNA-binding proteins called Cytoplasmic Polyadenylation Element-Binding Proteins (CPEB). If Orb2 is doing its job, the bug might instead have been reflecting on the one that got away at the moment of impact. 

As discussed in an interesting but rather technical article by Mujumdar A, et al., in drosophila (fruit flies) normal Orb2 forms degradation-resistant amyloid-like oligomers at neuronal synapses (in other words, functional amyloid in the connection points between brain nerve cells). These oligomers are essential to the persistence of long-term memory, which in this paper was determined by measuring "courtship suppression memory."  It goes something like this: a male fruit fly, after repeatedly being told to "buzz-off" by an uninterested female fruit fly, eventually learns to stop pestering her and other uninterested females. Normally, this newly-acquired behavior persists for days before waning. Although its tempting to discuss how eerily similar this sounds to a couple of desperate years in middle school, lets focus instead on what happens in fruit flies with mutant Orb2: within 24 hrs, they are back at it, pestering uninterested female fruit flies. A failure of long-term memory. 

Although CPEB is found in mammalian neurons too, and seems to play a role in memory formation, it isn't clear whether this is mediated by functional amyloid formation. As reviewed by J. Richter, stimulation of CPEB-containing nerve cells results in increases in specific neuronal proteins involved in memory persistence. Thus, it appears CPEB's function as a promotor of protein translation, rather than any amyloidogenic properties, may be the basis of its role in mammalian memory. Jury apparently still out, though. 

In the near future, I'll discuss another situation in which amyloid (amyloid-beta) plays a role in memory - in this case, the LOSS of memory - in people with Alzheimer's Dementia.    

21 February, 2013

Pomalidomide and Light Chain Amyloidosis

With the immunomodulatory drug pomalidomide (Pomalyst) recently approved for patients with multiple myeloma who have progressing disease within 60 days of the last of at least 2 prior therapies which had to have included lenalidomide (Revlimid) and bortezomib (Velcade), its an opportune time to review what we know (and don't know) about the drug in light chain (AL) amyloidosis. 

Pomalidomide, like lenalidomide, is a structural analogue of thalidomide. The structures of each are shown here:

source: http://www.readcube.com/articles/10.1186/2162-3619-1-27

The FDA-approved dose of POM in multiple myeloma is 4 mg/day for 21 out of every 28 days.

Investigators at the Mayo Clinic have studied POM (with dexamethasone) in 33 patients with previously treated AL amyloidosis. Patients had had a median of 2 prior therapies, with about half having had autologous stem cell transplant. A few (7) had previously been treated with other immunomodulatory drugs. 

The initial dose of POM in this study was 2 mg/day for 28 consecutive days each cycle (no break). DEX was given at 40 mg/week. POM dosing could be adjusted upward (if no response) and downward (for toxicity). The doses actually received are shown here: 

source: http://bloodjournal.hematologylibrary.org/content/119/23/5397.long

About half of the patients had hematologic responses, with the majority of these being partial. The median duration of response was 19 months. A handful of patients who had hematologic responses also had improvement in organ function, which is about what would be expected (based on prior work involving transplant, bortezomib, and other agents). Some of the responses observed occurred in patients who had had prior lenalidomide. 

A few notes regarding toxicity/adverse events: mild hematologic and gastrointestinal events were common. Peripheral sensory neuropathy (numbness, tingling) was documented in almost all patients - this was generally mild, but was more common than one might expect from, say, lenalidomide (at least in myeloma patients). The authors noted that the cardiac biomarker NT-pro-BNP (a marker of heart failure) sometimes worsened even in the setting of a hematologic response. Similar discordance between light chain measurements and cardiac markers have been noted previously in AL patients treated with immunomodulatory drugs (take a look).  In the present study, a 30% rise in the NT-pro-BNP in the first 3 months of therapy was associated with inferior survival, even though some such patients had concurrent light chain improvement.  

What we know: 
  • POM can (and undoubtedly will) be used as treatment for AL amyloidosis. 
  • Responses can be seen even in patients who have had prior autologous stem cell transplant, bortezomib, or lenalidomide.
What we don't know: 
  • Optimal dose, schedule, and duration of POM in AL amyloidosis (tsk! details!) 
  • Mechanism of NT-pro-BNP increase with POM therapy.
Study Spotlight: 

POM, like lenalidomide and thalidomide, will be studied in combination with other active drugs in patients with AL amyloidosis. The Karmanos Cancer Institute will be coordinating a national trial investigating the combination of POM, bortezomib, and dexamethasone in AL amyloidosis. This is the first trial prospectively studying an immunomodulatory drug combined with a proteasome inhibitor as front-line therapy for AL amyloidosis.  Participating centers will include Boston University, Duke University, the Colorado Blood Cancer Institute (Denver, CO) and Princess Margaret Hospital (Toronto).  

11 February, 2013

"Functional Amyloid"

"Functional amyloid" is a term I first heard about 3 years ago at the XII Amyloidosis Symposium in Rome. There was an oral presentation about the organization of peptide hormones in the secretory granules of the pituitary. Prior to that, I , like most clinicians, only thought about amyloid in the context of disease. It turns out amyloid actually plays an important role in vertebrate and invertebrate biology. 

In bacteria amyloid fibrils are involved in biofilm formation and cell/cell cell/substrate adhesion. Amyloid comprised of curli proteins in E.coli and Salmonella are a well-described example (see figure). Functional amyloid is also produced by Saccharomyces and Candida

source: http://www.mcdb.lsa.umich.edu/labs/chapman/research.php
The oval-shaped things are bacteria; the mesh-like stuff in between them is curli amyloid. 
As in amyloid diseases, the protein fibrils are extracellular; unlike these conditions, the production of amyloid appears to be a highly regulated process with intracellular trafficking, cellular export, fibril nucleation and the rapid polymerization requiring complex interactions between several curlin subunits (CsgA though G).  Its actually pretty interesting. No really, it is.

source: http://www.sciencedaily.com/releases/2008/02/080215121210.htm
A bacterial biofilm. Credit: Janice Haney Carr in Science Daily
In humans, a type of functional amyloid made up of a protein called pmel17 plays a role in the polymerization of the pigment melanin. Smaller melanin precursors (the building blocks from which melanin is ultimately constructed) would be toxic to the cell if they were not sequestered inside melanosomes by pmel17 amyloid. As defined by freemedicaldictionary.com, melanosomes are cigar- or oval-shaped pigment-containing granules within melanocytes. These granules are injected into keratinocytes, providing color:

source: http://www.katzenzeitung.eu/en/Genetics/melanocytes.html
Thus, pmel17 amyloid is critical for the formation of melanin and also cellular protection during this process. Click here for a link to an article discussing this in detail. 

The presentation in Rome I mentioned at the outset of this post discussed yet another functional role for amyloid in humans. Peptide hormones in the pituitary gland are actually stored as amyloid within secretory granules. It is postulated that this structure permits on the one hand, enduring stability in a highly concentrated state, and on the other, a means of controlled release of the hormones in question. Click here to read a publication from the authors of that presentation. 

There are illnesses characterized by abnormal amyloid or amyloid-like protein aggregates inside cells ("inclusion bodies"). An example would be Lewy Bodies in the neurons of patients with Parkinson's Disease. This type of protein accumulation is not functional amyloid, as it is generally damaging to the cell.

One can hope that understanding how cells handle functional intracellular amyloid will eventually provide insights into the pathophysiology of these disease states and perhaps lead to effective treatments.