Cysteine oxidation within N-terminal mutant huntingtin promotes oligomerization and delays clearance of soluble protein.

Huntington disease (HD) is a progressive neurodegenerative disorder caused by expression of polyglutamine-expanded mutant huntingtin protein (mhtt). Most evidence indicates that soluble mhtt species, rather than insoluble aggregates, are the important mediators of HD pathogenesis. However, the differential roles of soluble monomeric and oligomeric mhtt species in HD and the mechanisms of oligomer formation are not yet understood. We have shown previously that copper interacts with and oxidizes the polyglutamine-containing N171 fragment of huntingtin. In this study we report that oxidation-dependent oligomers of huntingtin form spontaneously in cell and mouse HD models. Levels of these species are modulated by copper, hydrogen peroxide, and glutathione. Mutagenesis of all cysteine residues within N171 blocks the formation of these oligomers. In cells, levels of oligomerization-blocked mutant N171 were decreased compared with native N171. We further show that a subset of the oligomerization-blocked form of glutamine-expanded N171 huntingtin is rapidly depleted from the soluble pool compared with "native " mutant N171. Taken together, our data indicate that huntingtin is subject to specific oxidations that are involved in the formation of stable oligomers and that also delay removal from the soluble pool. These findings show that inhibiting formation of oxidation-dependent huntingtin oligomers, or promoting their dissolution, may have protective effects in HD by decreasing the burden of soluble mutant huntingtin.

Mutations in amyloid precursor protein affect its interactions with presenilin/gamma-secretase.

Alzheimer's disease is characterized by accumulation of toxic beta-amyloid (Abeta) in the brain and neuronal death. Several mutations in presenilin (PS1) and beta-amyloid precursor protein (APP) associate with an increased Abeta(42/40) ratio. Abeta(42), a highly fibrillogenic species, is believed to drive Abeta aggregation. Factors shifting gamma-secretase cleavage of APP to produce Abeta(42) are unclear. We investigate the molecular mechanism underlying altered Abeta(42/40) ratios associated with APP mutations at codon 716 and 717. Using FRET-based fluorescence lifetime imaging to monitor APP-PS1 interactions, we show that I716F and V717I APP mutations increase the proportion of interacting molecules earlier in the secretory pathway, resulting in an increase in Abeta generation. A PS1 conformation assay reveals that, in the presence of mutant APP, PS1 adopts a conformation reminiscent of FAD-associated PS1 mutations, thus influencing APP binding to PS1/gamma-secretase. Mutant APP affects both intracellular location and efficiency of APP-PS1 interactions, thereby changing the Abeta(42/40) ratio.

N-cadherin-based adhesion enhances Abeta release and decreases Abeta42/40 ratio.

In neurons, Presenilin 1(PS1)/gamma-secretase is located at the synapses, bound to N-cadherin. We have previously reported that N-cadherin-mediated cell-cell contact promotes cell-surface expression of PS1/gamma-secretase. We postulated that N-cadherin-mediated trafficking of PS1 might impact synaptic PS1-amyloid precursor protein interactions and Abeta generation. In the present report, we evaluate the effect of N-cadherin-based contacts on Abeta production. We demonstrate that stable expression of N-cadherin in Chinese hamster ovary cells, expressing the Swedish mutant of human amyloid precursor protein leads to enhanced secretion of Abeta in the medium. Moreover, N-cadherin expression decreased Abeta(42/40) ratio. The effect of N-cadherin expression on Abeta production was accompanied by the enhanced accessibility of PS1/gamma-secretase to amyloid precursor protein as well as a conformational change of PS1, as demonstrated by the fluorescence lifetime imaging technique. These results indicate that N-cadherin-mediated synaptic adhesion may modulate Abeta secretion as well as the Abeta(42/40) ratio via PS1/N-cadherin interactions.

Cardiovascular Effects of the MEK Inhibitor, Trametinib: A Case Report, Literature Review, and Consideration of Mechanism.

The MEK inhibitor trametinib was approved in 2013 for the treatment of unresectable or metastatic melanoma with a BRAF V600E mutation, the most common pathogenic mutation in melanoma. Trametinib blocks activation of ERK1/2, inhibiting cell proliferation in melanoma. ERK1/2 also protects against multiple types of cardiac insult in mouse models. Trametinib improves survival in melanoma patients, but evidence of unanticipated cardiotoxicity is emerging. Here we describe the case of a patient with metastatic melanoma who developed acute systolic heart failure after trametinib treatment and present the results of the literature review prompted by this case. A patient with no cardiac history presented with a 6.5-mm skin lesion and was found to have metastatic BRAF V600E melanoma. Combination treatment with trametinib and the BRAF inhibitor, dabrafenib, was initiated. The patient's pre-treatment ejection fraction was 55-60%. His EF declined after 13 days and that was 40% 1 month after treatment. Two months after initiating trametinib, he developed dyspnea and fatigue. We conducted a chart review in the electronic medical record. We conducted a PubMed search using trametinib/adverse effects AND ("heart failure" OR "left ventricular dysfunction" OR hypertension OR cardiotoxicity OR mortality). We also queried the FDA Adverse Events Reporting System for reports of cardiomyopathy, ejection fraction decrease, and left ventricular dysfunction associated with trametinib between January 1, 2013, and July 20, 2017. The literature search retrieved 19 articles, including clinical trials and case reports. Early clinical experience with the MEK inhibitor trametinib suggests that its clinical efficacy may be compromised by cardiotoxicity. Further studies in humans and animals are required to determine the extent of this adverse effect, as well as its underlying mechanisms.

Applications for Research Concerning Fetal or Placental Tissue and Expected Institutional Review Board Responses.

Proposals for research concerning fetal and/or placental tissue may be refused institutional review board (IRB) review, effectively preventing the research from occurring. We conducted an anonymous electronic survey of IRB chairs to determine their assessment of the likely response to research projects using fetal/placental tissue obtained from various procedures. We found that proposals concerning tissue obtained from diagnostic procedures or miscarriage were anticipated to be considered at most institutions. Tissue obtained after abortion was likely to be refused consideration by more than 25% of respondents. Additional consultation during review was anticipated for up to 30% of scenarios. Responses for fetal and placental tissue were similar. The most frequently anticipated reason for refusal was institutional policy.

When the subject is more than just the subject: two case studies of family involvement in human subjects research.

Institutional review boards (IRBs) protect human research subjects by reviewing research to ensure compliance with federal regulations and institutional policies. One of the most important functions of IRBs is to ensure that investigators anticipate, plan for, and minimize risks to subjects. Under certain circumstances, however, participation in research may pose risks to nonsubject family members or other members of a subject's social network. In the context of a research protocol designed to test an intervention to prevent depression among a population of culturally diverse, urban mothers, we present two case studies of unanticipated problems, which demonstrate how nonsubject family members can either impact, or be impacted by, an individual's participation in research. The case studies illustrate the incongruence between federal regulations addressing IRB approval of research-which focus specifically on risks to subjects-and regulations on reporting incidents that occur during the conduct of the research, which extend to risks involving "others" as well. The cases also illustrate how risks to "others" can be accentuated in certain cultures where codependent family structures may increase the role that family members play in an individual's decision to participate in research. The question is raised as to whether this incongruence can inadvertently result in investigators and IRBs under-appreciating the risks that participation in research can pose to nonsubjects.

Variability in institutional approaches to ethics review of community-based research conducted in collaboration with unaffiliated organizations.

Academic institutions' requirements FOR ethics committee (IRB) review of research conducted by investigators from unaffiliated organizations engaged in collaborative, community-based research (CBR) may be highly variable. The present study examined the extent of this variability through a national survey of 196 IRB directors from US academic institutions. Fifty-three percent of respondents reported a formal policy or standardized approach to reviewing this type of CBR, with high volume IRBs more likely than low volume IRBs to do so (aOR 2.11, 95% CI 1.02, 4.35). The most common policy (40%) was to require that unaffiliated community organizations obtain a Federalwide Assurance on which they delegate responsibility for IRB review to their own (i.e., the academic institution's) IRB. Among IRBs without formal policies, 56% reported that human subject risk was their foremost consideration when reviewing CBR. Universities (71%) were more likely than medical schools (33%) to report subject risk as their foremost consideration (aOR 4.68, 95% CI 1.43, 15.28).