Characterization of the pathogenic α-Synuclein Variant V15A in Parkinson´s disease.

Despite being a major component of Lewy bodies and Lewy neurites, pathogenic variants in the gene encoding alpha-Synuclein (α-Syn) are rare. To date, only four missense variants in the SNCA gene, encoding α-Syn have unequivocally been shown to be disease-causing. We here describe a Parkinson´s disease patient with early cognitive decline carrying an as yet not fully characterized variant in SNCA (NM_001146055: c.44T > C, p.V15A). We used different cellular models, including stably transfected neuroblastoma (SH-SY5Y) cell cultures, induced pluripotent stem cell (iPSC)-derived neuronal cultures, and generated a Drosophila model to elucidate the impact of the p.V15A variant on α-Syn function and aggregation properties compared to other known pathogenic variants. We demonstrate that p.V15A increased the aggregation potential of α-Syn and the levels of apoptotic markers, and impaired the mitochondrial network. Moreover, p.V15A affects the flying ability and survival of mutant flies. Thus, we provide supporting evidence for the pathogenicity of the p.V15A variant, suggesting its inclusion in genetic testing approaches.

Differential Diagnosis and Molecular Stratification of Gastrointestinal Stromal Tumors on CT Images Using a Radiomics Approach.

Treatment planning of gastrointestinal stromal tumors (GISTs) includes distinguishing GISTs from other intra-abdominal tumors and GISTs' molecular analysis. The aim of this study was to evaluate radiomics for distinguishing GISTs from other intra-abdominal tumors, and in GISTs, predict the c-KIT, PDGFRA, BRAF mutational status, and mitotic index (MI). Patients diagnosed at the Erasmus MC between 2004 and 2017, with GIST or non-GIST intra-abdominal tumors and a contrast-enhanced venous-phase CT, were retrospectively included. Tumors were segmented, from which 564 image features were extracted. Prediction models were constructed using a combination of machine learning approaches. The evaluation was performed in a 100 × random-split cross-validation. Model performance was compared to that of three radiologists. One hundred twenty-five GISTs and 122 non-GISTs were included. The GIST vs. non-GIST radiomics model had a mean area under the curve (AUC) of 0.77. Three radiologists had an AUC of 0.69, 0.76, and 0.84, respectively. The radiomics model had an AUC of 0.52 for c-KIT, 0.56 for c-KIT exon 11, and 0.52 for the MI. The numbers of PDGFRA, BRAF, and other c-KIT mutations were too low for analysis. Our radiomics model was able to distinguish GISTs from non-GISTs with a performance similar to three radiologists, but less observer dependent. Therefore, it may aid in the early diagnosis of GIST, facilitating rapid referral to specialized treatment centers. As the model was not able to predict any genetic or molecular features, it cannot aid in treatment planning yet.

Incidence of unplanned excisions of soft tissue sarcomas in the Netherlands: A population-based study.

INTRODUCTION: Timely recognition of soft tissue sarcomas (STS) remains challenging, potentially leading to unplanned excisions (also known as 'whoops procedures'). This population-based study charted the occurrence of unplanned excisions and identified associated patient, tumour, and treatment-related characteristics. Furthermore, it presents an overview of the outcomes and clinical management following an unplanned excision. METHODS: From the Netherlands Cancer Registry (NCR) database, information was obtained on 2187 adult patients diagnosed with STS in 2016-2019 who underwent surgery. Tumours located in the mediastinum, heart or retroperitoneum were excluded, as well as incidental findings. Differences between patients with planned and unplanned excisions were assessed with chi-square tests and a multivariable logistic regression model. RESULTS: Overall, unplanned excisions comprise 18.2% of all first operations for STS, with a quarter of them occurring outside a hospital. Within hospitals, the unplanned excision rate was 14.4%. Unplanned excisions were more often performed on younger patients, and tumours unsuspected of being STS prior to surgery were generally smaller (≤5 cm) and superficially located. Preoperative imaging was omitted more frequently in these cases. An unplanned excision more often resulted in positive margins, requiring re-excision. Patients who had an unplanned excision outside of a sarcoma centre were more often discussed at or referred to a sarcoma centre, particularly in case of residual tumour. DISCUSSION: Potential improvement in preventing unplanned excisions may be achieved by better compliance to preoperative imaging and referral guidelines, and stimulating continuous awareness of STS among general surgeons, general practitioners and private practices.

MicroRNA expression and DNA methylation profiles do not distinguish between primary and recurrent well-differentiated liposarcoma.

Approximately one-third of the patients with well-differentiated liposarcoma (WDLPS) will develop a local recurrence. Not much is known about the molecular relationship between the primary tumor and the recurrent tumor, which is important to reveal potential drivers of recurrence. Here we investigated the biology of recurrent WDLPS by comparing paired primary and recurrent WDLPS using microRNA profiling and genome-wide DNA methylation analyses. In total, 27 paired primary and recurrent WDLPS formalin-fixed and paraffin-embedded tumor samples were collected. MicroRNA expression profiles were determined using TaqMan® Low Density Array (TLDA) cards. Genome-wide DNA methylation and differentially methylated regions (DMRs) were assessed by methylated DNA sequencing (MeD-seq). A supervised cluster analysis based on differentially expressed microRNAs between paired primary and recurrent WDLPS did not reveal a clear cluster pattern separating the primary from the recurrent tumors. The clustering was also not based on tumor localization, time to recurrence, age or status of the resection margins. Changes in DNA methylation between primary and recurrent tumors were extremely variable, and no consistent DNA methylation changes were found. As a result, a supervised clustering analysis based on DMRs between primary and recurrent tumors did not show a distinct cluster pattern based on any of the features. Subgroup analysis for tumors localized in the extremity or the retroperitoneum also did not yield a clear distinction between primary and recurrent WDLPS samples. In conclusion, microRNA expression profiles and DNA methylation profiles do not distinguish between primary and recurrent WDLPS and no putative common drivers could be identified.

Natural history of well-differentiated liposarcoma of the extremity compared to patients treated with surgery.

BACKGROUND: Patients with well-differentiated liposarcoma (WDLPS) of the extremity are mostly treated surgically, thereby possibly inducing severe morbidities. Despite the excellent prognosis, the natural history is barely studied. The aim of this study was to evaluate the natural history of extremity WDLPS by evaluating the outcome of patients treated with active surveillance (AS), who thereby exhibited the natural history of extremity WDLPS, and of patients treated surgically. METHODS: A large retrospective database of patients with extremity WDLPS was assessed to evaluate treatment, dedifferentiation and disease-specific survival. Lastly, our experience with patients treated with AS was explored. RESULTS: Distant metastases (5/191 patients, 2.6%) were mainly seen after a dedifferentiated local recurrence. Death of disease occurred in 4/191 patients (2.1%); two patients died from metastatic disease (although not pathologically proven), two patients died of treatment-related complications. In our center, 24 patients are treated with AS. Time of AS varied from 0.1 to 8.9 years (median 1.8). Four patients eventually underwent surgery after a period of AS (range 14-52 months) because of symptoms and/or tumor growth. No areas of dedifferentiation were found in these resection specimens. The other patients are still under active surveillance. CONCLUSION: Since surgical treatment might induce morbidity and even mortality, there might be overtreatment of these patients. Evaluation of the natural history of extremity WDLPS showed that AS could be a reasonable option for selected patients. Prospective studies in patients with extremity WDLPS are needed to assess the safety of AS as a treatment option.

Association of pazopanib-induced toxicities with outcome of patients with advanced soft tissue sarcoma; a retrospective analysis based on the European Organisation for Research and Treatment of Cancer (EORTC) 62043 and 62072 clinical trials.

Background: There is an unmet need for markers predicting the outcome of patients with advanced soft tissue sarcoma (STS) treated with pazopanib. Since toxicity might be related to the anti-tumor activity of the drug, the aim of this study was to determine whether pazopanib-induced proteinuria, hypothyroidism and cardiotoxicity grade 3-4 were associated with outcome. Methods: The combined results of the EORTC 62043 and 62072 trials were retrospectively assessed and used in a landmark analysis to evaluate the effect of the toxicities on progression-free survival (PFS) and overall survival (OS), using the Kaplan-Meier method and Cox regression models. Results: Of the 333 eligible patients, 259 patients were included in the analyses, for which a landmark time point of 60 days after randomization/registration was selected. Proteinuria occurred in 25.1%, hypothyroidism in 22.0% and cardiotoxicity grade 3-4 in 5.8% of the patients (any grade in 41.7%). There was no effect of the occurrence of proteinuria (6-months PFS 35.4% for patients with vs. 38.3% for patients without proteinuria, HR 1.01, p = .953), hypothyroidism (41.2% vs. 36.5%, HR 0.82, p = .210) or cardiotoxicity grade 3-4 (26.7% vs. 38.2%, HR 0.97, p = .897) on PFS. Nor was there an effect of proteinuria (6-months OS 63.2% for patients with vs. 74.4% for patients without proteinuria, HR 1.22, p = .196), hypothyroidism (76.2% vs. 70.5%, HR 0.75, p = .093) or cardiotoxicity grade 3-4 (80.0% vs. 77.2%, HR 0.93, p = .801) on OS. Conclusion: There was no association between the occurrence of pazopanib-induced proteinuria, hypothyroidism and cardiotoxicity and outcome. Therefore, these toxicities cannot be used as predictors for pazopanib activity in patients with advanced STS.

Increased survival of non low-grade and deep-seated soft tissue sarcoma after surgical management in high-volume hospitals: a nationwide study from the Netherlands.

BACKGROUND: Diagnosing and treating soft tissue sarcomas (STSs) remains challenging, stressing the urgency for centralisation. This nationwide survey aimed to evaluate the centralisation of STS surgery and its effect on survival. METHODS: Patients operated for primary STS from 2006 to 2015 were queried from the Netherlands Cancer Registry. Hospitals in which STS surgery was performed were allocated into three categories: low-volume (1-9 resections per year), medium-volume (10-19 resections) or high-volume (≥20 resections). Differences in tumour characteristics and outcome were calculated. A multivariable regression analysis was performed to adjust for case-mix. RESULTS: Of the 5282 identified patients, 42% was treated in low-volume hospitals, 7.7% in medium-volume hospitals and 51% in high-volume hospitals, with a significant trend over time towards treatment in a high-volume hospital (p < 0.01). In high-volume hospitals, more often patients with non low-grade, large and deep-seated tumours were treated than in low-volume hospitals. For the whole group, there was no survival benefit for patients treated in high-volume hospitals, with 10-year net survival rates of 76% (low-volume), 68% (medium-volume) and 68% (high-volume). However, subgroup analysis for patients with non low-grade and deep-seated tumours did reveal a benefit from treatment in a high-volume hospitals with 10-year survival rates of 54% (high-volume), 49% (low-volume) and 42% (medium-volume) and a relative risk of 1.3 (high-volume versus low-volume, p = 0.03). CONCLUSION: Centralisation of STS surgery has increased in the past decade. Surgery in a high-volume hospital improved survival of patients with non low-grade and deep-seated tumours, and therefore these patients should be referred to such a hospital.

Minimal Increase in Survival Throughout the Years in Patients with Soft Tissue Sarcoma with Synchronous Metastases: Results of a Population-Based Study.

BACKGROUND: Treatment options for patients with metastatic soft tissue sarcoma (STS) have increased in the last decade. We aimed to examine whether this is associated with improved overall survival (OS) in patients with STS with synchronous metastases. PATIENTS AND METHODS: Patients diagnosed with STS and synchronous metastases from 1989 to 2014 were queried from The Netherlands Cancer Registry. Trends in OS were assessed by the Kaplan-Meier method and log-rank test in time intervals of 5 years, for the whole study population and in subgroups for liposarcomas, leiomyosarcoma, and other STS subtypes. A multivariable Cox regression analysis was performed to identify characteristics prognostic for OS. RESULTS: Median OS of the 1,393 identified patients did not improve significantly over the years from 5.8 months in 1989-1994 to 8.1 months in 2010-2014, but there was an evident trend. Median OS was prolonged in the subgroups of liposarcomas (3.6 to 9.3 months), leiomyosarcomas (11.3 to 14.6 months), and other STS subtypes (5.7 to 6.3 months), although there were no significant improvements in OS over the years. Primary tumor site in one of the extremities and surgery in an academic center had a favorable effect on OS, whereas significant negative predictors were no treatment, elderly age, STS subtype other than liposarcoma or leiomyosarcoma, high or unknown grade, and nodal involvement. CONCLUSION: Although overall survival of patients with STS with synchronous metastases in this nationwide and "real-life" population has improved over the years, the improvement was not statistically significant, despite new treatment options. IMPLICATIONS FOR PRACTICE: Treatment of patients with metastatic soft tissue sarcoma (STS) has changed in the past years, with new drugs such as trabectedin (2007) and pazopanib (2012) becoming available. By using data from the nationwide Netherlands Cancer Registry, the impact of these changes in treatment policies on survival is analyzed in a "real-life" population of patients with STS with synchronous metastases, rather than in a strictly selected trial population. Unfortunately, overall survival improved only minimally and not significantly for these patients diagnosed from 1989 to 2014. Hopefully, the advent of novel treatment options, such as eribulin and olaratumab, will further improve the outcome of this patient group.

EJC's biennial report on metastatic soft tissue sarcoma: State of the art and future perspectives.

In the last decade the limited treatment options for patients with metastatic soft tissue sarcoma have expanded considerably. With the addition of olaratumab to first-line treatment with doxorubicin, the introduction of several new agents in second-line treatment and beyond and other promising agents in the pipeline, perspectives of patients with metastatic soft tissue sarcoma are improving. Due to increasing insight into the biology of the different soft tissue sarcoma subtypes, choice of treatment has become much more histology-driven, although more prognostic and predictive factors are needed to further personalise therapy. This report summarises the current state of the art and discusses the promising developments in the treatment of patients with metastatic soft tissue sarcoma.

Cardiolipin promotes electron transport between ubiquinone and complex I to rescue PINK1 deficiency.

PINK1 is mutated in Parkinson's disease (PD), and mutations cause mitochondrial defects that include inefficient electron transport between complex I and ubiquinone. Neurodegeneration is also connected to changes in lipid homeostasis, but how these are related to PINK1-induced mitochondrial dysfunction is unknown. Based on an unbiased genetic screen, we found that partial genetic and pharmacological inhibition of fatty acid synthase (FASN) suppresses toxicity induced by PINK1 deficiency in flies, mouse cells, patient-derived fibroblasts, and induced pluripotent stem cell-derived dopaminergic neurons. Lower FASN activity in PINK1 mutants decreases palmitate levels and increases the levels of cardiolipin (CL), a mitochondrial inner membrane-specific lipid. Direct supplementation of CL to isolated mitochondria not only rescues the PINK1-induced complex I defects but also rescues the inefficient electron transfer between complex I and ubiquinone in specific mutants. Our data indicate that genetic or pharmacologic inhibition of FASN to increase CL levels bypasses the enzymatic defects at complex I in a PD model.

Near-infrared 808 nm light boosts complex IV-dependent respiration and rescues a Parkinson-related pink1 model.

Mitochondrial electron transport chain (ETC) defects are observed in Parkinson's disease (PD) patients and in PD fly- and mouse-models; however it remains to be tested if acute improvement of ETC function alleviates PD-relevant defects. We tested the hypothesis that 808 nm infrared light that effectively penetrates tissues rescues pink1 mutants. We show that irradiating isolated fly or mouse mitochondria with 808 nm light that is absorbed by ETC-Complex IV acutely improves Complex IV-dependent oxygen consumption and ATP production, a feature that is wavelength-specific. Irradiating Drosophila pink1 mutants using a single dose of 808 nm light results in a rescue of major systemic and mitochondrial defects. Time-course experiments indicate mitochondrial membrane potential defects are rescued prior to mitochondrial morphological defects, also in dopaminergic neurons, suggesting mitochondrial functional defects precede mitochondrial swelling. Thus, our data indicate that improvement of mitochondrial function using infrared light stimulation is a viable strategy to alleviate pink1-related defects.

Aconitase causes iron toxicity in Drosophila pink1 mutants.

The PTEN-induced kinase 1 (PINK1) is a mitochondrial kinase, and pink1 mutations cause early onset Parkinson's disease (PD) in humans. Loss of pink1 in Drosophila leads to defects in mitochondrial function, and genetic data suggest that another PD-related gene product, Parkin, acts with pink1 to regulate the clearance of dysfunctional mitochondria (mitophagy). Consequently, pink1 mutants show an accumulation of morphologically abnormal mitochondria, but it is unclear if other factors are involved in pink1 function in vivo and contribute to the mitochondrial morphological defects seen in specific cell types in pink1 mutants. To explore the molecular mechanisms of pink1 function, we performed a genetic modifier screen in Drosophila and identified aconitase (acon) as a dominant suppressor of pink1. Acon localizes to mitochondria and harbors a labile iron-sulfur [4Fe-4S] cluster that can scavenge superoxide to release hydrogen peroxide and iron that combine to produce hydroxyl radicals. Using Acon enzymatic mutants, and expression of mitoferritin that scavenges free iron, we show that [4Fe-4S] cluster inactivation, as a result of increased superoxide in pink1 mutants, results in oxidative stress and mitochondrial swelling. We show that [4Fe-4S] inactivation acts downstream of pink1 in a pathway that affects mitochondrial morphology, but acts independently of parkin. Thus our data indicate that superoxide-dependent [4Fe-4S] inactivation defines a potential pathogenic cascade that acts independent of mitophagy and links iron toxicity to mitochondrial failure in a PD-relevant model.

Vitamin K2 is a mitochondrial electron carrier that rescues pink1 deficiency.

Human UBIAD1 localizes to mitochondria and converts vitamin K(1) to vitamin K(2). Vitamin K(2) is best known as a cofactor in blood coagulation, but in bacteria it is a membrane-bound electron carrier. Whether vitamin K(2) exerts a similar carrier function in eukaryotic cells is unknown. We identified Drosophila UBIAD1/Heix as a modifier of pink1, a gene mutated in Parkinson's disease that affects mitochondrial function. We found that vitamin K(2) was necessary and sufficient to transfer electrons in Drosophila mitochondria. Heix mutants showed severe mitochondrial defects that were rescued by vitamin K(2), and, similar to ubiquinone, vitamin K(2) transferred electrons in Drosophila mitochondria, resulting in more efficient adenosine triphosphate (ATP) production. Thus, mitochondrial dysfunction was rescued by vitamin K(2) that serves as a mitochondrial electron carrier, helping to maintain normal ATP production.

Genomic characterization of novel dolphin papillomaviruses provides indications for recombination within the Papillomaviridae.

Phylogenetic analysis of novel dolphin (Tursiops truncatus) papillomavirus sequences, TtPV1, -2, and -3, indicates that the early and late protein coding regions of their genomes differ in evolutionary history. Sliding window bootscan analysis showed a significant a change in phylogenetic clustering, in which the grouped sequences of TtPV1 and -3 move from a cluster with the Phocoena spinipinnis PsPV1 in the early region to a cluster with TtPV2 in the late region. This provides indications for a possible recombination event near the end of E2/beginning of L2. A second possible recombination site could be located near the end of L1, in the upstream regulatory region. Selection analysis by using maximum likelihood models of codon substitutions ruled out the possibility of intense selective pressure, acting asymmetrically on the viral genomes, as an alternative explanation for the observed difference in evolutionary history between the early and late genomic regions of these cetacean papillomaviruses.