By: Lise Munsie, PhD
In the lab…
The Ranum laboratory published their discovery of Repeat Associated Non-ATG (RAN) translation in HD in Neuron.1 This was the first demonstration of RAN translation in a disease where the causative triplet repeat expansion occurs in the coding region. They showed that poly-Ala, poly-Cys, poly-Leu and poly-Ser proteins are translated in neurons that express huntingtin (Htt) mini-genes with disease-causing CAG expansion lengths. Accumulation of these proteins increases with expansion size. They found RAN-translated proteins in brain regions affected by disease in post-mortem HD brain tissue, with abundance and protein specificity correlating with disease severity.
Bowles and colleagues used the immortalized striatal cell line derived from Q111 mice to examine phosphorylation of Htt by stimulating the epidermal growth factor (EGF) pathway.2 They found alterations in MEK and AKT phosphorylation of wild-type compared to mutant Htt (mHtt) after EGF stimulation, leading to altered nuclear localization and altered transcription of a subset of genes. Inhibiting these kinases in the presence of EGF affects Htt localization, especially in the mutant cell line. The authors hypothesize it may be possible to attenuate these changes using MEK and AKT inhibitors. Kinase inhibitors may correct Htt localization, attenuating pathology associated with altered transcription.
Whether over-expression or activation of 5′ adenosine monophosphate-activated protein kinase (AMPK) is neuroprotective or toxic in HD remains controversial. Using nematode models and murine striatal neuron models, a recent report showed that AMPK over-expression has positive outcomes in models.3 Using fluorescence resonance energy transfer (FRET) techniques, Vázquez-Manrique and colleagues showed a decrease in soluble mHtt in the presence of AMPK activation. In vivo, they demonstrated that active AMPK co-transfected with toxic Htt species results in smaller brain lesions, but no alteration in the number of ubiquitin-positive aggregates.
2Bowles KR, Brooks SP, Dunnett SB, Jones L. Huntingtin subcellular localisation is regulated by kinase signalling activity in the StHdhQ111 model of HD. PloS one. 2015;10(12):e0144864.
3Vázquez-Manrique RP, Farina F, Cambon K, et al. AMPK activation protects from neuronal dysfunction and vulnerability across nematode, cellular and mouse models of Huntington’s disease. Hum Mol Genet. 2015 Dec 17. pii: ddv513. [Epub ahead of print].
In the pipeline…
Using an unbiased screen for huntingtin (HTT)-interacting transcription factors, the La Spada group identified peroxisome proliferator-activated receptor delta (PPAR-δ) as a novel HTT interactor.1 An alteration in PPAR-δ transactivation in a polyglutamine-dependent manner leads to mitochondrial dysfunction, enhancing cell death. When injected into the central nervous system of mice, a dominant-negative PPAR construct produced motor dysfunction, neurodegeneration, and mitochondrial abnormalities consistent with HD. Pharmacologically inducing PPAR-δ activity ameliorated HD-induced dysfunction and cell death in mouse and stem cell striatal cell models. The authors propose that KD3010, a compound which enhances PPAR-δ activation, may be useful as a treatment for aspects of HD pathology.
The Messer laboratory performed studies examining the safety of active vaccination as a treatment for HD in mice.2 They assessed the safety and immunogenicity of 11 immunization protocols, including different combinations of peptide, protein and DNA vaccines. They found that a combination of three non-overlapping Htt exon-1 peptides resulted in the greatest immunogenicity, but also led to transcriptional dysregulation of immune-related genes. This treatment strategy may necessitate adjuvant therapies aimed at restoring immune controls.
Finding heterozygous polymorphisms that exist in cis with the mHTT allele is essential to development of allele-specific silencing therapies. A recent report in Molecular Therapy prioritized target selection by examining diverse HD patient populations across Canada, France, Sweden, and Italy.3 The group defined three target haplotypes, A1, A2, and A3, that could be targeted by allele-specific antisense oligonucleotides (ASOs) to treat the 80% of patients who have one of these haplotypes. The group demonstrated that ASOs against polymorphisms in the A1 haplotypes selectively inhibit mHTT in a potent and specific manner.
1Dickey AS, Pineda VV, Tsunemi T, et al. PPAR-[delta] is repressed in Huntington’s disease, is required for normal neuronal function and can be targeted therapeutically. Nat Med. 2015 Dec 7. doi: 10.1038/nm.4003. [Epub ahead of print].
2Ramsingh AI, Manley K, Rong Y, et al. Transcriptional dysregulation of inflammatory/immune pathways after active vaccination against Huntington′s disease. Hum Mol Genet. 2015 Nov 1;24(21):6186-97. doi: 10.1093/hmg/ddv335. Epub 2015 Aug 24.
3Kay C, Collins JA, Skotte NH, et al. Huntingtin haplotypes provide prioritized target panels for allele-specific silencing in Huntington disease patients of European ancestry. Mol Ther. 2015;23(11):1759-1771.
In stem cells…
The Ellerby group created isogenic induced pluripotent stem cell (iPSC) lines from HD patients’ cells.1 In a recent Stem Cell report, the group performed transcriptome analysis on the HD and the isogenic lines. They found a 10-fold increase in differentially expressed genes in the HD line compared to the isogenic line after differentiation to neural stem cells, compared to differentially expressed genes prior to differentiation. The group showed that transforming growth factor (TGF-β) and netrin-1 signaling pathways are differentially regulated at the neural stem cell stage, and can be modulated to rescue HD-related phenotypes in both the in vitro stem cell model and an in vivo mouse model.
Another recent report utilized iPSCs derived from the YAC128 mouse model, an adult-onset HD patient, and a juvenile-onset HD patient, to investigate pathways involved in HD pathogenesis.2 Multiple signaling pathways were altered in these lines, including the MAPK/ERK pathways, and p53 signaling. Other pathways (such as TGF-β) were not affected in the undifferentiated cells. This paper indicates that some, but not all, pathways are affected early and, possibly, peripherally in HD.
Stimulation of the A2A adenosine receptor (A2AR) in cell culture models of HD has positive anti-apoptotic outcomes. A paper published in Human Molecular Genetics used iPSCs derived from HD and control patients differentiated into DARPP32 GABAergic neurons, and examined how stimulating A2AR differentially effects the two cell lines.3 Treating the cells with hydrogen peroxide introduced oxidative stress and caused double-stranded DNA breaks, with more double-stranded breaks in the HD-derived neurons. A2AR agonists ameliorated this phenotype in the differentiated neurons through PKA activation. The authors suggest the use of this platform for drug screening.
1Ring Karen L, An Mahru C, Zhang N, et al. Genomic analysis reveals disruption of striatal neuronal development and therapeutic targets in human Huntington’s disease neural stem cells. Stem Cell Reports. 2015 Dec 8;5(6):1023-38. doi: 10.1016/j.stemcr.2015.11.005.
2Szlachcic WJ, Switonski PM, Krzyzosiak WJ, et al. Huntington disease iPSCs show early molecular changes in intracellular signaling, the expression of oxidative stress proteins and the p53 pathway. Dis Model Mech. 2015 Sep;8(9):1047-57. doi: 10.1242/dmm.019406. Epub 2015 Jun 18.
3Chiu F-L, Lin J-T, Chuang C-Y, et al. Elucidating the role of the A2A adenosine receptor in neurodegeneration using neurons derived from Huntington’s disease iPSCs. Hum Mol Genet. 2015 Nov 1;24(21):6066-79. doi: 10.1093/hmg/ddv318. Epub 2015 Aug 11.