Impaired mitochondrial morphological plasticity and failure of mitophagy associated with the G11778A mutation of LHON.

Mitochondrial dynamics were examined in human dermal fibroblasts biopsied from a confirmed Leber's Hereditary Optic Neuropathy (LHON) patient with a homoplasmic G11778A mutation of the mitochondrial genome. Expression of the G11778A mutation did not impart any discernible difference in mitochondrial network morphology using widefield fluorescence microscopy. However, at the ultrastructural level, cells expressing this mutation exhibited an impairment of mitochondrial morphological plasticity when forced to utilize oxidative phosphorylation (OXPHOS) by transition to glucose-free, galactose-containing media. LHON fibroblasts also displayed a transient increase in mitophagy upon transition to galactose media. These results provide new insights into the consequences of the G11778A mutation of LHON and the pathological mechanisms underlying this disease.

Meaningful Symptomatic Change in Patients With Myelofibrosis From the SIMPLIFY Studies.

OBJECTIVES: Patients with myelofibrosis develop symptoms due to bone marrow fibrosis, systemic inflammation, and/or organomegaly. Alleviating symptoms improves overall quality of life. Clinical trials have historically defined symptom response as a reduction of at least 50% in Total Symptom Score at week 24 compared with baseline. Whether 50% constitutes a meaningful benefit has not been established. This study determined the meaningful change threshold (MCT) for 2 momelotinib phase III trials, SIMPLIFY-1 and SIMPLIFY-2. METHODS: The absolute and percentage MCT was determined using anchor-based methods applied to the modified Myeloproliferative Neoplasm Symptom Assessment Form v2.0 and Patient Global Impression of Change. MCTs were applied retrospectively to determine responder rates. Generalized estimating equations estimated the treatment-related difference in likelihood of improvement. RESULTS: In SIMPLIFY-1, a Janus kinase inhibitor-naive population, the MCT was 8 points. In SIMPLIFY-2, a previously Janus kinase inhibitor-treated population, the MCT was 6 points. A 32% MCT was determined in both studies, showing that the historic 50% reduction threshold may be a conservative choice. In SIMPLIFY-1, a similar proportion of patients achieved responder status with 24 weeks of momelotinib or ruxolitinib therapy based on the absolute MCT (39% vs 41%, respectively). In SIMPLIFY-2, a significantly greater proportion of patients treated with momelotinib achieved responder states compared with best available therapy based on absolute and percent change MCTs. CONCLUSIONS: This study demonstrates that momelotinib provided clinically meaningful symptom benefit for patients with myelofibrosis and provides insight into the appropriateness of the symptom change threshold used in historical studies.

Changes in mitochondrial morphology induced by calcium or rotenone in primary astrocytes occur predominantly through ros-mediated remodeling.

Morphological changes in mitochondria have been primarily attributed to fission and fusion, while the more pliable transformations of mitochondria (remodeling, rounding, or stretching) have been largely overlooked. In this study, we quantify the contributions of fission and remodeling to changes in mitochondrial morphology induced by the Ca(2+) ionophore 4Br-A23187 and the metabolic toxin rotenone. We also examine the role of reactive oxygen species (ROS) in the regulation of mitochondrial remodeling. In agreement with our previous studies, mitochondrial remodeling, not fission, is the primary contributor to Ca(2+) -mediated changes in mitochondrial morphology induced by 4Br-A23187 in rat cortical astrocytes. Treatment with rotenone produced similar results. In both paradigms, remodeling was selectively blocked by antioxidants whereas fission was not, suggesting a ROS-mediated mechanism for mitochondrial remodeling. In support of this hypothesis, inhibition of endogenous ROS by overnight incubation in antioxidants resulted in elongated reticular networks of mitochondria. Examination of inner and outer mitochondrial membranes revealed that they largely acted in concert during the remodeling process. While mitochondrial morphology is traditionally ascribed to a net output of fission and fusion processes, in this study we provide evidence that the acute pliability of mitochondria can be a dominant factor in determining their morphology. More importantly, our results suggest that the remodeling process is independently regulated through a ROS-signaling mechanism. Mitochondrial morphology is traditionally ascribed to a balance of fission and fusion processes. We have shown that mitochondria can undergo more pliable transformations; remodeling, rounding, or stretching. We demonstrate that remodeling, not fission, is the primary contributor to calcium mediated changes in mitochondrial morphology in primary astrocytes. Others have shown fission is mediated by calcineurin. Our results suggest the remodeling process distinct from fission and is independently regulated through a ROS-signaling mechanism (CsA: Cyclosporine A; NAC: N-acetyl-l-cysteine; GSH: Reduced-L-Glutathione).

The C. elegans CDK8 Mediator module regulates axon guidance decisions in the ventral nerve cord and during dorsal axon navigation.

Receptors expressed on the growth cone of outgrowing axons detect cues required for proper navigation. The pathway choices available to an axon are in part defined by the set of guidance receptors present on the growth cone. Regulated expression of receptors and genes controlling the localization and activity of receptors ensures that axons respond only to guidance cues relevant for reaching their targets. In genetic screens for axon guidance mutants, we isolated an allele of let-19/mdt-13, a component of the Mediator, a large ~30 subunit protein complex essential for gene transcription by RNA polymerase II. LET-19/MDT-13 is part of the CDK8 module of the Mediator. By testing other Mediator components, we found that all subunits of the CDK8 module as well as some other Mediator components are required for specific axon navigation decisions in a subset of neurons. Expression profiling demonstrated that let-19/mdt-13 regulates the expression of a large number of genes in interneurons. A mutation in the sax-3 gene, encoding a receptor for the repulsive guidance cue SLT-1, suppresses the commissure navigation defects found in cdk-8 mutants. This suggests that the CDK8 module specifically represses the SAX-3/ROBO pathway to ensure proper commissure navigation.

Mitochondrial trafficking in neuropsychiatric diseases.

Mitochondria have numerous roles in healthy neuronal functioning and in neuronal injury mechanisms. They are quite dynamic organelles in that they fuse, divide and move throughout axons and dendrites. The mechanisms of mitochondrial motility have received much attention, however the significance of the dynamic nature of mitochondria in neurons is unclear. Nonetheless, deficits in mitochondrial trafficking have been implicated in numerous neurodegenerative disorders. The role of aberrant mitochondrial trafficking in neuropsychiatric disorders is not as well understood, but may involve similar mechanisms. In this review we examine the evidence which implicates changes in mitochondrial trafficking in the pathogenesis of neuropsychiatric disorders and hypothesize how defective mitochondrial transport may contribute to disease mechanisms.

Symptomatic benefit of momelotinib in patients with myelofibrosis: Results from the SIMPLIFY phase III studies.

BACKGROUND: Myelofibrosis (MF)-associated constitutional symptoms can severely impact health-related quality of life. Clinical trials in MF traditionally measure symptom response to treatment as a landmark endpoint of total symptom score (TSS) reduction ≥50% from baseline. However, this dichotomous assessment provides a limited view of clinically relevant symptomatic changes. Herein we evaluated longitudinal change from baseline in TSS over the continuous 24-week period and individual symptom scores to obtain a more comprehensive understanding of symptom benefits experienced by patients with MF receiving therapy. METHODS: Longitudinal symptom change was evaluated using mixed-effect model repeated measure (MMRM) methodology with individual item-level analyses to complement the interpretation of the landmark symptom results in the completed phase III SIMPLIFY studies of momelotinib in MF. MMRM compared mean change in TSS from baseline with Week 24 using data from all patient visits. Generalized estimating equations were used to estimate item-level odds ratios using multiple predictive imputations for missing data. RESULTS: Momelotinib and ruxolitinib groups reported similar overall symptom improvements, with a TSS difference of <1.5 points between groups for each post-baseline visit in SIMPLIFY-1. In SIMPLIFY-2, the improvement in TSS observed in momelotinib-treated patients was consistent with that observed in SIMPLIFY-1, whereas progressive TSS deterioration was observed with control. Item-level scores were heterogeneous in both studies. A similar and greater proportion of momelotinib-treated patients were categorized as "improved" or "stable" compared with control in SIMPLIFY-1 and SIMPLIFY-2, respectively. Odds ratios for between-group comparison ranged from 0.75 to 1.21 in SIMPLIFY-1, demonstrating similarity in likelihood of symptom improvement. In SIMPLIFY-2, the likelihood of symptom improvement in each item was higher in the momelotinib arm. CONCLUSIONS: These findings suggest that momelotinib provides clinically relevant symptom benefits in the JAK inhibitor-naïve and JAK inhibitor-exposed settings.

Patient-reported Outcomes and Quality of Life in Anemic and Symptomatic Patients With Myelofibrosis: Results From the MOMENTUM Study.

Myelofibrosis (MF) is a chronic myeloproliferative neoplasm that typically manifests with debilitating symptoms that progressively worsen, negatively impacting patients' quality of life. Fatigue is a multifactorial and burdensome MF-related symptom due to its severity, persistence, and prevalence, with anemia a contributing factor and major unmet need. Clinical trials of the Janus kinase (JAK)1/JAK2/activin A receptor type 1 inhibitor momelotinib have shown consistent anemia benefits, in addition to improvements in MF-related symptoms. The phase 3 MOMENTUM trial in symptomatic and anemic patients met its primary end point, with a greater proportion having a Myelofibrosis Symptom Assessment Form (MFSAF) Total Symptom Score (TSS) reduction ≥50% at week 24 with momelotinib versus danazol. To support the positive primary end point result, we conducted longitudinal, responder, and time-to-event analyses of patient-reported outcomes from MOMENTUM, as measured by the MFSAF, European Organization for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-C30), and Patient-Reported Outcomes Measurement Information System (PROMIS) assessments. These analyses demonstrated rapid and durable response benefits with momelotinib, with achievement of first TSS response by day 29 and continued improvement over time. Improvements favored momelotinib versus danazol for each MFSAF individual item, and greater improvements were observed for disease- and cancer-related fatigue and physical functioning at week 24, with significant results for multiple items/domains across the 3 assessments. These findings are consistent in demonstrating that momelotinib provides substantial symptom benefit.

Genomic Classifier for Guiding Treatment of Intermediate-Risk Prostate Cancers to Dose-Escalated Image Guided Radiation Therapy Without Hormone Therapy.

PURPOSE: The National Comprehensive Cancer Network (NCCN) has recently endorsed the stratification of intermediate-risk prostate cancer (IR-PCa) into favorable and unfavorable subgroups and recommend the addition of androgen deprivation therapy (ADT) to radiation therapy (RT) for unfavorable IR-PCa. Recently, more accurate prognostication was demonstrated by integrating a 22-feature genomic classifier (GC) to the NCCN stratification system. Here, we test the utility of the GC to better identify patients with IR-PCa who are sufficiently treated by RT alone. METHODS AND MATERIALS: We identified a novel cohort comprising 121 patients with IR-PCa treated with dose-escalated image guided RT (78 Gy in 39 fractions) without ADT. GC scores were derived from tumors sampled in diagnostic biopsies. Multivariable analyses, including both NCCN subclassification and GC scores, were performed for biochemical failure (prostate-specific antigen nadir + 2 ng/mL) and metastasis occurrence. RESULTS: By NCCN subclassification, 33 (27.3%) and 87 (71.9%) of men were classified as having favorable and unfavorable IR-PCa, respectively (1 case unclassifiable). GC scores were high in 3 favorable IR-PCa and low in 60 unfavorable IR-PCa. Higher GC scores, but not NCCN risk subgroups, were associated with biochemical relapse (hazard ratio, 1.36; 95% confidence interval [CI], 1.09-1.71] per 10% increase; P = .007) and metastasis (hazard ratio, 2.05; 95% CI, 1.24-4.24; P = .004). GC predicted biochemical failure at 5 years (area under the curve, 0.78; 95% CI, 0.59-0.91), and the combinatorial NCCN + GC model significantly outperformed the NCCN alone model for predicting early-onset metastasis (area under the curve for 5-year metastasis of 0.89 vs 0.86 [GC alone] vs 0.54 [NCCN alone]). CONCLUSIONS: We demonstrated the accuracy of the GC for predicting disease recurrence in IR-PCa treated with dose-escalated image guided RT alone. Our findings highlight the need to evaluate this GC in a prospective clinical trial investigating the role of ADT-RT in clinicogenomic-defined IR-PCa subgroups.

Validation of the Decipher Test for predicting adverse pathology in candidates for prostate cancer active surveillance.

ABSTACT: BACKGROUND: Many men diagnosed with prostate cancer are active surveillance (AS) candidates. However, AS may be associated with increased risk of disease progression and metastasis due to delayed therapy. Genomic classifiers, e.g., Decipher, may allow better risk-stratify newly diagnosed prostate cancers for AS. METHODS: Decipher was initially assessed in a prospective cohort of prostatectomies to explore the correlation with clinically meaningful biologic characteristics and then assessed in diagnostic biopsies from a retrospective multicenter cohort of 266 men with National Comprehensive Cancer Network (NCCN) very low/low and favorable-intermediate risk prostate cancer. Decipher and Cancer of the Prostate Risk Assessment (CAPRA) were compared as predictors of adverse pathology (AP) for which there is universal agreement that patients with long life-expectancy are not suitable candidates for AS (primary pattern 4 or 5, advanced local stage [pT3b or greater] or lymph node involvement). RESULTS: Decipher from prostatectomies was significantly associated with adverse pathologic features (p-values < 0.001). Decipher from the 266 diagnostic biopsies (64.7% NCCN-very-low/low and 35.3% favorable-intermediate) was an independent predictor of AP (odds ratio 1.29 per 10% increase, 95% confidence interval [CI] 1.03-1.61, p-value 0.025) when adjusting for CAPRA. CAPRA area under curve (AUC) was 0.57, (95% CI 0.47-0.68). Adding Decipher to CAPRA increased the AUC to 0.65 (95% CI 0.58-0.70). NPV, which determines the degree of confidence in the absence of AP for patients, was 91% (95% CI 87-94%) and 96% (95% CI 90-99%) for Decipher thresholds of 0.45 and 0.2, respectively. Using a threshold of 0.2, Decipher was a significant predictor of AP when adjusting for CAPRA (p-value 0.016). CONCLUSION: Decipher can be applied to prostate biopsies from NCCN-very-low/low and favorable-intermediate risk patients to predict absence of adverse pathologic features. These patients are predicted to be good candidates for active surveillance.

Ability of a Genomic Classifier to Predict Metastasis and Prostate Cancer-specific Mortality after Radiation or Surgery based on Needle Biopsy Specimens.

BACKGROUND: Decipher is a validated genomic classifier developed to determine the biological potential for metastasis after radical prostatectomy (RP). OBJECTIVE: To evaluate the ability of biopsy Decipher to predict metastasis and Prostate cancer-specific mortality (PCSM) in primarily intermediate- to high-risk patients treated with RP or radiation therapy (RT). DESIGN, SETTING, AND PARTICIPANTS: Two hundred and thirty-five patients treated with either RP (n=105) or RT±androgen deprivation therapy (n=130) with available genomic expression profiles generated from diagnostic biopsy specimens from seven tertiary referral centers. The highest-grade core was sampled and Decipher was calculated based on a locked random forest model. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Metastasis and PCSM were the primary and secondary outcomes of the study, respectively. Cox analysis and c-index were used to evaluate the performance of Decipher. RESULTS AND LIMITATIONS: With a median follow-up of 6 yr among censored patients, 34 patients developed metastases and 11 died of prostate cancer. On multivariable analysis, biopsy Decipher remained a significant predictor of metastasis (hazard ratio: 1.37 per 10% increase in score, 95% confidence interval [CI]: 1.06-1.78, p=0.018) after adjusting for clinical variables. For predicting metastasis 5-yr post-biopsy, Cancer of the Prostate Risk Assessment score had a c-index of 0.60 (95% CI: 0.50-0.69), while Cancer of the Prostate Risk Assessment plus biopsy Decipher had a c-index of 0.71 (95% CI: 0.60-0.82). National Comprehensive Cancer Network risk group had a c-index of 0.66 (95% CI: 0.53-0.77), while National Comprehensive Cancer Network plus biopsy Decipher had a c-index of 0.74 (95% CI: 0.66-0.82). Biopsy Decipher was a significant predictor of PCSM (hazard ratio: 1.57 per 10% increase in score, 95% CI: 1.03-2.48, p=0.037), with a 5-yr PCSM rate of 0%, 0%, and 9.4% for Decipher low, intermediate, and high, respectively. CONCLUSIONS: Biopsy Decipher predicted metastasis and PCSM from diagnostic biopsy specimens of primarily intermediate- and high-risk men treated with first-line RT or RP. PATIENT SUMMARY: Biopsy Decipher predicted metastasis and prostate cancer-specific mortality risk from diagnostic biopsy specimens.

Individual Patient-Level Meta-Analysis of the Performance of the Decipher Genomic Classifier in High-Risk Men After Prostatectomy to Predict Development of Metastatic Disease.

Purpose To perform the first meta-analysis of the performance of the genomic classifier test, Decipher, in men with prostate cancer postprostatectomy. Methods MEDLINE, EMBASE, and the Decipher genomic resource information database were searched for published reports between 2011 and 2016 of men treated by prostatectomy that assessed the benefit of the Decipher test. Multivariable Cox proportional hazards models fit to individual patient data were performed; meta-analyses were conducted by pooling the study-specific hazard ratios (HRs) using random-effects modeling. Extent of heterogeneity between studies was determined with the I2 test. Results Five studies (975 total patients, and 855 patients with individual patient-level data) were eligible for analysis, with a median follow-up of 8 years. Of the total cohort, 60.9%, 22.6%, and 16.5% of patients were classified by Decipher as low, intermediate, and high risk, respectively. The 10-year cumulative incidence metastases rates were 5.5%, 15.0%, and 26.7% ( P < .001), respectively, for the three risk classifications. Pooling the study-specific Decipher HRs across the five studies resulted in an HR of 1.52 (95% CI, 1.39 to 1.67; I2 = 0%) per 0.1 unit. In multivariable analysis of individual patient data, adjusting for clinicopathologic variables, Decipher remained a statistically significant predictor of metastasis (HR, 1.30; 95% CI, 1.14 to 1.47; P < .001) per 0.1 unit. The C-index for 10-year distant metastasis of the clinical model alone was 0.76; this increased to 0.81 with inclusion of Decipher. Conclusion The genomic classifier test, Decipher, can independently improve prognostication of patients postprostatectomy, as well as within nearly all clinicopathologic, demographic, and treatment subgroups. Future study of how to best incorporate genomic testing in clinical decision-making and subsequent treatment recommendations is warranted.

Elevated intracellular calcium causes distinct mitochondrial remodelling and calcineurin-dependent fission in astrocytes.

Disruptions of mitochondrial dynamics have been implicated in the pathogenesis of neurodegenerative diseases. The regulation mechanisms of mitochondrial dynamics have not been fully elucidated; however, calcium has been suggested to play a role. In the present study, we examined the role of intracellular calcium in regulating mitochondrial morphology and motility in cortical astrocytes employing different concentrations of a calcium ionophore. High levels of calcium caused a dramatic reduction in mitochondrial length, the result of two distinct phenomena: mitochondrial remodelling (or "rounding") and fission. Quantitative analysis revealed that mitochondrial remodelling/rounding was the predominant process. In addition, mitochondrial motility was reduced, as reported previously in neurons. By contrast, prolonged, more modest levels of intracellular calcium resulted in a reduction in mitochondrial length without significant effects upon mitochondrial motility. This calcium-induced reduction in mitochondrial length was not affected by the presence of calcineurin inhibitors; however, when mitochondrial fission events were specifically examined, calcineurin inhibitors had a significant inhibitory effect. This suggests that changes in mitochondrial length were primarily due to mitochondrial remodelling as opposed to fission. In the present study, we have therefore dissected the effects of calcium on mitochondrial motility, remodelling and fission. Our results suggest independent mechanisms for regulating these processes.