What a failed drug does (and is there hope for latrepirdine?)

Latrepirdine (Dimebon) was initially used as an antihistamine drug in Russia. It was later found to be neuroprotective, and entered phase II clinical trials in the US for both Alzheimer’s disease and Huntington’s disease. However, Dimebon failed in a US-based phase II replication trial of a prior successful Russian phase II trial of mild-to-moderate AD. Given the initial promise of the drug and split results,  as well as the lack of treatments for neurodegenerative diseases, there in is significant interest in understanding the underlying molecular mechanism(s) for the drug’s effects.

In a paper appearing this week in Molecular Psychiatry, Brandeis researchers in the Petsko-Ringe lab, including postdoc Shulin Ju and undergraduate Jessica Liken ’11, used yeast models of neurodegenerative disease associated proteins to show that Dimebon specifically protects yeast from the cytotoxiciy of α-synuclein, a protein involved in Parkinson’s disease. They further showed that protection is mediated through its up-regulation of autophagy pathway. In collaboration with Sam Gandy‘s group at Mount Sinai School of Medicine, these findings were further confirmed and validated in neuronal cell and animal models.

Given these observations, disparities in the contribution of α-synuclein to the neuropathology between the Russian and US Dimebon studies might also explain, at least in part, the inconsistency of the cognitive benefit in the two trials. If this speculation is correct, then it may be interesting to test for benefits of Dimebon in treating synucleinopathies such as Parkinson’s disease, Lewy body dementia, REM sleep disorder and/or multiple system atrophy.

see also: press release from Mt. Sinai Alzheimer’s Diesease Research Center

Steele JW (*), Ju S(*), Lachenmayer ML(*), Liken J, Stock A, Kim SH, Delgado LM, Alfaro IE, Bernales S, Verdile G, Bharadwaj P, Gupta V, Barr R, Friss A, Dolios G, Wang R, Ringe D, Protter AA, Martins RN, Ehrlich ME, Yue Z, Petsko GA, Gandy S. Latrepirdine stimulates autophagy and reduces accumulation of alpha-synuclein in cells and in mouse brain. Molecular psychiatry. 2012.

Steele JW(*), Lachenmayer ML(*), Ju S, Stock A, Liken J, Kim SH, Delgado LM, Alfaro IE, Bernales S, Verdile G, Bharadwaj P, Gupta V, Barr R, Friss A, Dolios G, Wang R, Ringe D, Fraser P, Westaway D, St George-Hyslop PH, Szabo P, Relkin NR, Buxbaum JD, Glabe CG, Protter AA, Martins RN, Ehrlich ME, Petsko GA, Yue Z, Gandy S. Latrepirdine improves cognition and arrests progression of neuropathology in an Alzheimer’s mouse model. Molecular psychiatry. 2012.

Dynamics of double-strand break repair

In a new paper in the journal Genetics, former Brandeis postdoc Eric Coïc and undergrads Taehyun Ryu and Sue Yen Tay from Professor of Biology Jim Haber’s lab, along with grad student Joshua Martin and Professor of Physics Jané Kondev, tackle the problem of understanding the dynamics of homologous recombination after double strand breaks in yeast. According to Haber,

The accurate repair of chromosome breaks is an essential process that prevents cells from undergoing gross chromosomal rearrangements that are the hallmark of most cancer cells.  We know a lot about how such breaks are repaired.  The ends of the break are resected and provide a platform for the assembly of many copies of the key recombination protein, Rad51.  Somehow the Rad51 filament is then able to facilitate a search of the entire DNA of the nucleus to locate identical or nearly identical (homologous) sequences so that the broken end can pair up with this template and initiate local copying of this segment to patch up the chromosome break.  How this search takes place remains poorly understood.

The switching of budding yeast mating type genes has been a valuable model system in which to study the molecular events of broken chromosome repair, in real time.  It is possible to induce synchronously a site-specific double-strand break (DSB) on one chromosome, within the mating-type (MAT) locus.  At opposite ends of the same chromosome are two competing donor sequences with which the broken ends of the MAT sequence can pair up and copy new mating-type sequences into the MAT locus.

Normally one of these donors is used 9 times more often than the other.  We asked if this preference was irrevocable or if the bias could be changed by making the “wrong” donor more attractive – in this case by adding more sequences to that donor so that it shared more and more homology with the broken ends at MAT.  We found that the competition could indeed be changed and that adding more homologous sequences to the poorly-used donor increased its use.

In collaboration with Jané Kondev’s lab we devised both a “toy” model and a more rigorous thermodynamic model to explain these results.  They suggest that the Rad51 filament carrying the broken end of the MAT locus collides on average 4 times before with the preferred donor region before it actually succeeds in carrying out the next steps in the process that lead to repair and MAT switching.

Dynamics of homology searching during gene conversion in Saccharomyces cerevisiae revealed by donor competition Eric Coïc , Joshua Martin, Taehyun Ryu, Sue Yen Tay, Jané Kondev and James E. Haber. Genetics. 2011 Sep 27 2011 Sep 27

Phosphatases and DNA double strand break repair

When cells suffer DNA damage – as little as a single break in one chromosome – they respond by activating the DNA damage checkpoint, which prevents cells from entering mitosis until there is enough time to to repair the damage.  The principal biochemical events in the checkpoint pathway are the phosphorylations of protein kinases by other protein kinases and eventually the phosphorylation of other proteins that regulate mitosis.    When repair is complete, the checkpoint must be turned off.  Not surprisingly, the enzymes that turn off the checkpoint are phosphatases that can remove the phosphates added by the protein kinases.

The Haber lab has previously shown that, in budding yeast, a pair of PP2C phosphatases known as Ptc2 and Ptc3 were important in turning off a key protein kinase, Rad53.  A member of another phosphatase subgroup, the PP4 phosphatase Pph3, dephosphorylates a target of the checkpoint kinases, histone protein H2A.  There is one aspect that they didn’t understand at all: It seems that the intensity of the checkpoint signals must grow the longer it takes to repair DNA damage, because deletions of ptc2 and ptc3 or a deletion of pph3 prevented cells from turning off the damage signal when it took a long time – 6 hours – to repair the damage, but they had much less effect on different repair events that could complete in 3-4 hours or in less than 2 hours.  So they decided to see what would happen if they created a yeast strain lacking all three phosphatases (ptc2 ptc3 pph3), leading to a paper appearing this month in the journal Molecular and Cell Biology.

To their surprise, these cells had a new defect: they couldn’t complete the repair event itself, rather than simply being defective in resuming mitosis after repair was completed.  The mutants could not properly initiate the small amounts of DNA copying that are required for repair.  Again, the severity of the defect depends on the length of the delay it takes to initiate the repair event itself.  The figure (right) shows that the triple mutant is also much more sensitive to DNA damaging agents such as the anti-cancer drug camptothecin (CPT) and to methylmethansulfonate (MMS). These data show a complex connection between DNA damage signaling and the repair process itself, and reveal new roles for the phosphatases in DNA repair.  The work was carried out primarily by graduate student Jung-Ae Kim, now a postdoc at Rockefeller University, with help by another grad student, Wade Hicks, and by an undergraduate Sue Yen Tay, and postdoc Jin Li. The work was supported by a research and a graduate student training grant from the NIH.

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