Readsynth: short-read simulation for consideration of composition-biases in reduced metagenome sequencing approaches.

BACKGROUND: The application of reduced metagenomic sequencing approaches holds promise as a middle ground between targeted amplicon sequencing and whole metagenome sequencing approaches but has not been widely adopted as a technique. A major barrier to adoption is the lack of read simulation software built to handle characteristic features of these novel approaches. Reduced metagenomic sequencing (RMS) produces unique patterns of fragmentation per genome that are sensitive to restriction enzyme choice, and the non-uniform size selection of these fragments may introduce novel challenges to taxonomic assignment as well as relative abundance estimates. RESULTS: Through the development and application of simulation software, readsynth, we compare simulated metagenomic sequencing libraries with existing RMS data to assess the influence of multiple library preparation and sequencing steps on downstream analytical results. Based on read depth per position, readsynth achieved 0.79 Pearson's correlation and 0.94 Spearman's correlation to these benchmarks. Application of a novel estimation approach, fixed length taxonomic ratios, improved quantification accuracy of simulated human gut microbial communities when compared to estimates of mean or median coverage. CONCLUSIONS: We investigate the possible strengths and weaknesses of applying the RMS technique to profiling microbial communities via simulations with readsynth. The choice of restriction enzymes and size selection steps in library prep are non-trivial decisions that bias downstream profiling and quantification. The simulations investigated in this study illustrate the possible limits of preparing metagenomic libraries with a reduced representation sequencing approach, but also allow for the development of strategies for producing and handling the sequence data produced by this promising application.

Olfactory bulb anomalies in KBG syndrome mouse model and patients.

ANKRD11 (ankyrin repeat domain 11) is a chromatin regulator and the only gene associated with KBG syndrome, a rare neurodevelopmental disorder. We have previously shown that Ankrd11 regulates murine embryonic cortical neurogenesis. Here, we show a novel olfactory bulb phenotype in a KBG syndrome mouse model and two diagnosed patients. Conditional knockout of Ankrd11 in murine embryonic neural stem cells leads to aberrant postnatal olfactory bulb development and reduced size due to reduction of the olfactory bulb granule cell layer. We further show that the rostral migratory stream has incomplete migration of neuroblasts, reduced cell proliferation as well as aberrant differentiation of neurons. This leads to reduced neuroblasts and neurons in the olfactory bulb granule cell layer. In vitro, Ankrd11-deficient neural stem cells from the postnatal subventricular zone display reduced migration, proliferation, and neurogenesis. Finally, we describe two clinically and molecularly confirmed KBG syndrome patients with anosmia and olfactory bulb and groove hypo-dysgenesis/agenesis. Our report provides evidence that Ankrd11 is a novel regulator of olfactory bulb development and neuroblast migration. Moreover, our study highlights a novel clinical sign of KBG syndrome linked to ANKRD11 perturbations in mice and humans.

Clinical variables influencing the perception of fatigue in people with multiple sclerosis: a cross-sectional study using FSIQ-RMS.

BACKGROUND: Physical fatigue is one of the most disabling symptoms in people with Multiple Sclerosis (PwMS). Several factors might influence the development of fatigue, such as gender, education, body mass index (BMI), Expanded Disability Status Scale (EDSS), disease duration, working status (Ws), physiotherapy (Ph), and disease-modifying therapies (DMTs). Fatigue Symptoms and Impacts Questionnaire-Relapsing Multiple Sclerosis (FSIQ-RMS) is a patient-reported outcome (PRO) that allows one to define the impact of fatigue in PwMS clearly. This study aimed to assess fatigue impact on PwMS by using FSIQ-RMS. METHODS: The participants were enrolled from May to July 2021 in MS Centers of Sant'Andrea Hospital and Policlinico Umberto I Hospital in Rome. Fatigue was evaluated using the FSIQ-RMS, validated, and culturally adapted in Italian. Clinical and demographic data were collected at the same time. RESULTS: We enrolled 178 PwMS [Female 74.16%; RMS 82.58%, SPMS 17.52%]. FSIQ-RMS scores were significantly correlated with EDSS (p-value < 0.01). Analysis of variance between means showed a statistically significant difference between the BMI groups at the 24hours_FSIQ-RMS score and the 7days_FSIQ-RMS score (p < 0.01), with the lower BMI group having the highest scores. Furthermore, perceived fatigue significantly improved both in subjects performing Ph (p < 0.05) and in those who actively work (p < 0.01). CONCLUSIONS: The use of FSIQ-RMS in a real-world setting confirmed that underweight and high levels of disability are closely related to fatigue. In addition, Ph and active Ws are strongly correlated with fatigue in PwMS.

Synthesis of two Fluorescent Complexes and Their use as Multifunctional Nanomedicine Carriers for Rhabdomyosarcoma Treatment.

This study focuses on the design and synthesis of two novel coordination polymers (CPs), named 1 and 2, with excellent fluorescent properties. Their structures were characterized by X-ray single-crystal diffraction, revealing that both materials exhibit promising fluorescence performance, indicating their potential as fluorescent detection tools. Additionally, 1 was chosen to be combined with chitosan (CS), resulting in the successful fabrication of a biodegradable and non-toxic efficient drug carrier, termed CS-1@Cisplatin. This carrier possesses a large surface area and good solubility, enabling sustained drug release to target cells. Given that CXC motif chemokine receptor type 4 (CXCR4) is a key marker gene highly expressed in Rhabdomyosarcoma (RMS) cells and tissues, RMS was chosen as the biological model for testing. The results demonstrated that CS-1@Cisplatin effectively inhibited the invasiveness of RMS cells by significantly suppressing CXCR4 expression. Therefore, the system shows great potential for applications in RMS treatment, biometrics, and drug delivery, particularly in its unique advantage of targeting RMS by inhibiting the key marker gene CXCR4.

The mechanisms underpinning the slow component of [Formula: see text] in humans.

PURPOSE: When exercising above the lactic threshold (LT), the slow component of oxygen uptake ([Formula: see text]) appears, mainly ascribed to the progressive recruitment of Type II fibers. However, also the progressive decay of the economy of contraction may contribute to it. We investigated oxygen uptake ([Formula: see text]) during isometric contractions clamping torque (T) or muscular activation to quantify the contributions of the two mechanisms. METHODS: We assessed for 7 min T of the leg extensors, net oxygen uptake ([Formula: see text]) and root mean square (RMS) from vastus lateralis (VL) in 11 volunteers (21 ± 2 yy; 1.73 ± 0.11 m; 67 ± 14 kg) during cyclic isometric contractions (contraction/relaxation 5 s/5 s): (i) at 65% of maximal voluntary contraction (MVC) (FB-Torque) and; (ii) keeping the level of RMS equal to that at 65% of MVC (FB-EMG). RESULTS: [Formula: see text] after the third minute in FB-Torque increased with time ([Formula: see text] = 94 × t + 564; R2 = 0.99; P = 0.001), but not during FB-EMG. [Formula: see text]/T increased only during FB-Torque ([Formula: see text]/T = 1.10 × t + 0.57; R2 = 0.99; P = 0.001). RMS was larger in FB-Torque than in FB-EMG and significantly increased in the first three minutes of exercise to stabilize till the end of the trial, indicating that the pool of recruited MUs remained constant despite [Formula: see text]. CONCLUSION: The analysis of the RMS, [Formula: see text] and T during FB-Torque suggests that the intrinsic mechanism attributable to the decay of contraction efficiency was responsible for an increase of [Formula: see text] equal to 18% of the total [Formula: see text].

Complete mimicry: Rhabdomyosarcoma with FUS::TFCP2 fusion masquerading as carcinoma-diagnostic challenge and report of two cases.

Rhabdomyosarcomas (RMS) are malignant mesenchymal tumors with skeletal muscle differentiation which are classified into alveolar, embryonal, pleomorphic, and spindle cell/sclerosing RMS. Within the spindle cell/sclerosing RMS tumor type there is a recently recognized sub-type categorized as intraosseous spindle cell RMS with TFCP2/NCOA2 gene fusion. This rare tumor is highly aggressive with predominant involvement of the craniofacial and pelvic bones with approximately 30 cases reported to date. Histopathologic features include spindle cell and epithelioid morphology with a characteristic co-expression of epithelial markers, myogenic markers, and ALK1 expression. We report two cases of gnathic spindle cell/sclerosing RMS with FUS::TFCP2 gene fusion that were initially interpreted as carcinomas by referring institutions and later reclassified when encountered in our practice after additional work-up and molecular characterization.

Changes on chromosome 11p15.5 as specific marker for embryonal rhabdomyosarcoma?

Rhabdomyosarcomas (RMS) constitute a heterogeneous spectrum of tumors with respect to clinical behavior and tumor morphology. The paternal uniparental disomy (pUPD) of 11p15.5 is a molecular change described mainly in embryonal RMS. In addition to LOH, UPD, the MLPA technique (ME030kit) also determines copy number variants and methylation of H19 and KCNQ1OT1 genes, which have not been systematically investigated in RMS. All 127 RMS tumors were divided by histology and PAX status into four groups, pleomorphic histology (n = 2); alveolar RMS PAX fusion-positive (PAX+; n = 39); embryonal RMS (n = 70) and fusion-negative RMS with alveolar pattern (PAX-RMS-AP; n = 16). The following changes were detected; negative (n = 21), pUPD (n = 75), gain of paternal allele (n = 9), loss of maternal allele (n = 9), hypermethylation of H19 (n = 6), hypomethylation of KCNQ1OT1 (n = 6), and deletion of CDKN1C (n = 1). We have shown no difference in the frequency of pUPD 11p15.5 in all groups. Thus, we have proven that changes in the 11p15.5 are not only specific to the embryonal RMS (ERMS), but are often also present in alveolar RMS (ARMS). We have found changes that have not yet been described in RMS. We also demonstrated new potential diagnostic markers for ERMS (paternal duplication and UPD of whole chromosome 11) and for ARMS PAX+ (hypomethylation KCNQ1OT1).

Development of carbon monoxide-releasing molecules conjugated to polysaccharides (glyco-CORMs) for delivering CO during obesity.

Metal carbonyls have been developed as carbon monoxide-releasing molecules (CO-RMs) to deliver CO for therapeutic purposes. The manganese-based CORM-401 has been recently reported to exert beneficial effects in obese animals by reducing body weight gain, improving glucose metabolism and reprogramming adipose tissue towards a healthy phenotype. Here, we report on the synthesis and characterization of glyco-CORMs, obtained by grafting manganese carbonyls on dextrans (70 and 40 kDa), based on the fact that polysaccharides facilitate the targeting of drugs to adipose tissue. We found that glyco-CORMs efficiently deliver CO to cells in vitro with higher CO accumulation in adipocytes compared to other cell types. Oral administration of two selected glyco-CORMs (5b and 6b) resulted in CO accumulation in various organs, including adipose tissue. In addition, glyco-CORM 6b administered for eight weeks elicited anti-obesity and positive metabolic effects in mice fed a high fat diet. Our study highlights the feasibility of creating carriers with multiple functionalized CO-RMs.

Rhabdomyosarcoma xenotransplants in zebrafish embryos.

Rhabdomyosarcomas (RMS) are the most common pediatric soft tissue sarcomas. High-risk and metastatic disease continues to be associated with very poor prognosis. RMS model systems that faithfully recapitulate the human disease and provide rapid, cost-efficient estimates of antitumor efficacy of candidate drugs are needed to facilitate drug development and personalized medicine approaches. Here, we present a new zebrafish-based xenotransplant model allowing for rapid and easily accessible drug screening using low numbers of viable tumor cells and relatively small amounts of water-soluble chemicals. Under optimized temperature conditions, embryonal RMS xenografts were established in zebrafish embryos at 3 h postfertilization (hpf). In proof-of-principle experiments, chemotherapy drugs with established clinical anti-RMS efficacy (vincristine, dactinomycin) and the mitogen-activated protein kinase kinase inhibitor trametinib were shown to significantly reduce the cross-sectional area of the tumors by 120 hpf. RMS xenograft models in zebrafish embryos henceforth could serve as a valuable addition to cell culture and mammalian models of RMS and represent a rapid and cost-effective solution for preclinical candidate drug testing.

Micro-relief characterization of benign and malignant skin lesions by polarization speckle analysis in vivo.

BACKGROUND/PURPOSE: A recent direction in skin disease classification is to develop quantitative diagnostic techniques. Skin relief, colloquially known as roughness, is an important clinical feature. The aim of this study is to demonstrate a novel polarization speckle technique to quantitatively measure roughness on skin lesions in vivo. We then calculate the average roughness of different types of skin lesions to determine the extent to which polarization speckle roughness measurements can be used to identify skin cancer. METHODS: The experimental conditions were set to target the fine relief structure on the order of ten microns within a small field of view of 3 mm. The device was tested in a clinical study on patients with malignant and benign skin lesions that resemble cancer. The cancer group includes 37 malignant melanomas (MM), 43 basal cell carcinomas (BCC), and 26 squamous cell carcinomas (SCC), all categories confirmed by gold standard biopsy. The benign group includes 109 seborrheic keratoses (SK), 79 nevi, and 11 actinic keratoses (AK). Normal skin roughness was obtained for the same patients (301 different body sites proximal to the lesion). RESULTS: The average root mean squared (rms) roughness ± standard error of the mean for MM and nevus was equal to 19 ± 5 µm and 21 ± 3 µm, respectively. Normal skin has rms roughness of 31 ± 3 µm, other lesions have roughness of 35 ± 10 µm (AK), 35 ± 7 µm (SCC), 31 ± 4 µm (SK), and 30 ± 5 µm (BCC). CONCLUSION: An independent-samples Kruskal-Wallis test indicates that MM and nevus can be separated from each of the tested types of lesions, except each other. These results quantify clinical knowledge of lesion roughness and could be useful for optical cancer detection.

Comparing criteria for deviation from hearing aid prescriptive targets in children.

OBJECTIVE: To determine if a stricter criterion for paediatric hearing aid fitting for proximity of fit-to-target of <3 dB root-mean-square (RMS) error produces better audibility and outcomes compared to the current <5 dB criterion, and to examine the relationship between aided audibility and RMS error by degree of hearing loss. DESIGN: We evaluated the influence of unaided hearing level on the relationship between RMS error and aided audibility. We assessed the effect of RMS error category (<3, 3-5, >5 dB) on aided audibility, speech recognition, expressive vocabulary, and morphosyntax. STUDY SAMPLE: The study included 2314 hearing aid verification measurements from 307 children with hearing aids. RESULTS: Children who met a <3 dB criterion had higher aided audibility than children who met no criterion (>5 dB error). Results showed no differences in speech recognition or vocabulary by error category, but children with <3 dB error demonstrated better morphosyntax than children with 3-5 and >5 dB RMS error. CONCLUSIONS: Fittings that are close to prescriptive targets provide a more positive outcome for children with hearing aids. Using probe microphone measures to adjust hearing aids to within 3 dB may benefit language abilities in children.

The Actual Exactness of a Fast RMS Correction during Abrupt Voltage Change.

The requirement of RMS (voltage and current) measurements under a fraction of the AC period has become increasingly attractive in power systems. Some of these power applications are responsible for voltage stabilization in distribution lines when the voltage correction should be made in a short time, no more than one or two periods of the AC signal. Previously developed RMS correction applications must be validated in real-world situations characterized by an abrupt change (discontinuity) in voltage magnitude occurring even during a single AC period. Such circumstances can substantially influence the RMS estimation and, therefore, should be considered. This article suggests a mathematically based approach, validated in the laboratory, that improves the accuracy of a voltage RMS estimation for the appropriate measurement devices. It produces better results in cases where the RMS assessment should be done in a fraction of the AC period.

A Study on the Effect of Temperature Variations on FPGA-Based Multi-Channel Time-to-Digital Converters.

We describe a study on the effect of temperature variations on multi-channel time-to-digital converters (TDCs). The objective is to study the impact of ambient thermal variations on the performance of field-programmable gate array (FPGA)-based tapped delay line (TDL) TDC systems while simultaneously meeting the requirements of high-precision time measurement, low-cost implementation, small size, and low power consumption. For our study, we chose two devices, Artix-7 and ProASIC3L, manufactured by Xilinx and Microsemi, respectively. The radiation-tolerant ProASIC3L device offers better stability in terms of thermal sensitivity and power consumption compared to the Artix-7. To assess the performance of the TDCs under varying thermal conditions, a laboratory thermal chamber was utilized to maintain ambient temperatures ranging from -75 to 80 °C. This analysis ensured a comprehensive evaluation of the TDCs' performance across a wide operational range. By utilizing the Artix-7 and ProASIC3L devices, we achieved root mean square (RMS) resolution of 24.7 and 554.59 picoseconds, respectively. Total on-chip power of 0.968 W was achieved using Artix-7, while 1.997 mW of power consumption was achieved using the ProASIC3L device. We worked to determine the temperature sensitivity for both FPGA devices, which could help in the design and optimization of FPGA-based TDCs for many applications.

Garcinol and Anacardic Acid, Natural Inhibitors of Histone Acetyltransferases, Inhibit Rhabdomyosarcoma Growth and Proliferation.

Rhabdomyosarcoma (RMS) is a malignant tumour of the soft tissues. There are two main histopathological types: alveolar and embryonal. RMS occurs mainly in childhood and is a result of the deregulation of growth and differentiation of muscle cell precursors. There is an increasing amount of data indicating that numerous epigenetic alterations within chromatin and histone proteins are involved in the pathogenesis of this malignancy. Histone acetylation is one of the most important epigenetic modifications that is catalysed by enzymes from the group of histone acetyltransferases (HAT). In this study, the impact of the natural histone acetyltransferase inhibitors (HATi)-garcinol (GAR) and anacardic acid (AA)-on the biology of RMS cells was evaluated through a series of in vitro tests measuring proliferation, viability, clonogenicity, cell cycle and apoptosis. Moreover, using oligonucleotide microarrays and real-time PCR, we identified several genes whose expression changed after GAR and AA treatment. The examined HATi significantly reduce the invasive phenotype of RMS cells by inhibiting the growth rate, viability and clonogenic abilities. What is more, these substances cause cell cycle arrest in the G2/M phase, induce apoptosis and affect the genetic expression of the endoplasmic reticulum stress sensors. GAR and AA may serve as promising potential anti-cancer drugs since they sensitize the RMS cells to chemotherapeutic treatment.

Assessing the impact of land use and land cover on river water quality using water quality index and remote sensing techniques.

The impact of land use on water quality is becoming a global concern due to the increasing demand for freshwater. This study aimed to assess the effects of land use and land cover (LULC) on the surface water quality of the Buriganga, Dhaleshwari, Meghna, and Padma river system in Bangladesh. To determine the state of water, water samples were collected from twelve locations in the Buriganga, Dhaleshwari, Meghna, and Padma rivers during the winter season of 2015 and collected samples were analysed for seven water quality indicators: pH, temperature (Temp.), conductivity (Cond.), dissolved oxygen (DO), biological oxygen demand (BOD), nitrate nitrogen (NO3-N), and soluble reactive phosphorus (SRP) for assessing water quality (WQ). Additionally, same-period satellite imagery (Landsat-8) was utilised to classify the LULC using the object-based image analysis (OBIA) technique. The overall accuracy assessment and kappa co-efficient value of post-classified images were 92% and 0.89, respectively. In this research, the root mean squared water quality index (RMS-WQI) model was used to determine the WQ status, and satellite imagery was utilised to classify LULC types. Most of the WQs were found within the ECR guideline level for surface water. The RMS-WQI result showed that the "fair" status of water quality found in all sampling sites ranges from 66.50 to 79.08, and the water quality is satisfactory. Four types of LULC were categorised in the study area mainly comprised of agricultural land (37.33%), followed by built-up area (24.76%), vegetation (9.5%), and water bodies (28.41%). Finally, the Principal component analysis (PCA) techniques were used to find out significant WQ indicators and the correlation matrix revealed that WQ had a substantial positive correlation with agricultural land (r = 0.68, P < 0.01) and a significant negative association with the built-up area (r = - 0.94, P < 0.01). To the best of the authors' knowledge, this is the first attempt in Bangladesh to assess the impact of LULC on the water quality along the longitudinal gradient of a vast river system. Hence, we believe that the findings of this study can support planners and environmentalists to plan and design landscapes and protect the river environment.

SIX1 as a Novel Immunohistochemical Marker in the Differential Diagnosis of Rhabdomyosarcoma.

Background: Differential diagnosis of rhabdomyosarcoma (RMS) is challenging. Sineoculis homeobox homolog 1 (SIX1) is an oncogene involved in skeletal muscle differentiation. We compared protein expression patterns of SIX1 in RMS and its most common differential diagnoses. Methods: SIX1 immunohistochemistry in 36 RMS and in 33 tumors from seven differential diagnostic subtypes were evaluated. The fraction of SIX1 positive tumor cells was scored by three independent observers. Results: A majority (75%) of the evaluated RMS expressed SIX1 in at least 50% of tumor cells and all except one RMS had more than 25% positive tumor cells. Neuroblastoma had less than 1% SIX1 positive tumor cells. Gonadoblastoma, malignant rhabdoid tumor, and Ewing sarcoma had 10% or less positive tumor cells. Pleuropulmonary blastoma exhibited 26-50% positive tumor cells and synovial sarcoma >50% positive cells. Conclusion: SIX1 immunohistochemistry is positive in most RMS, and occasionally in some tumors within the differential diagnoses of RMS.

The effects of additive manufacturing technologies and finish line designs on the trueness and dimensional stability of 3D-printed dies.

PURPOSE: To evaluate the effects of 5 manufacturing technologies and 2 finish line designs on the trueness and dimensional stability of 3D-printed definitive dies at finish line regions under different storage conditions and time. MATERIAL AND METHODS: Preparation of light chamfer and round shoulder finish lines were adopted individually on two mandibular first molar typodont teeth and digitalized as standard tessellation language (STL) files. A total of 240 samples (192 AM definitive dies and 48 definitive conventional stone dies) in 20 groups (n = 12) were manufactured based on 2 finishing line designs (chamfer and shoulder), 5 manufacturing technologies (4 additively manufactured technologies and conventional stone die), and 2 storage conditions (light exposure and dark). The 4 additively manufactured (AM) technologies include a DLP 3D-printer, an economic LED 3D-printer, a CLIP 3D-printer, and an SLA 3D-printer. All the study samples were distributed into two storage conditions. Subsequently, samples were digitalized to STL files at 3 different time points (within 36 hours, 1-month, and 3-months). A surface matching software was used to superimpose the sample STL files onto the corresponding original STL files with the best-fit alignment function. The trueness of each printed and stone definitive dies and their dimensional stabilities were measured by the root mean square (RMS, in mm). A linear mixed-effects model was used to test the effects of the finish line design, manufacturing technology, storage condition, and storage time on RMS values (α = 0.05). RESULTS: While finish line designs had no significant effects [F(1, 220) = 0.85, p < 0.358], the manufacturing technologies [F(3, 220) = 33.02, p < 0.001], storage condition [F(1, 220) = 4.11, p = 0.044], and storage time F(2, 440) = 10.37, p < 0.001] affected the trueness and dimensional stability of 3D-printed dies at finish line regions. No significant interactions were found among the 4 factors. For the manufacturing technologies, Type IV stone groups and LCD 3D-printer groups had significantly higher RMS values than the other 3 printers (p < 0.001) with no significant differences between Type IV stone and LCD 3D-printer groups (p = 0.577). DLP 3D-printer groups had higher RMS values than both SLA 3D-printer groups and CLIP 3D-printer groups (p < 0.001). There were no significant differences between SLA 3D-printer groups and CLIP 3D-printer groups, p = 0.671. For the effects of storage conditions, RMS values were significantly higher in the groups stored with the direct light exposure than the ones stored in the dark, p = 0.044. In terms of the effects of storage time, the RMS values were significantly higher after 1-month storage, p = 0.002; and 3-month storage, p < 0.001, than the ones at the immediate postmanufacturing stage. However, the RMS values after 1-month and 3-month storage were not significantly different from each other (p = 0.169). CONCLUSIONS: Manufacturing technologies, storage conditions, and storage time significantly affected the trueness and dimensional stability of 3D-printed dies at finish line regions, while finish line designs had no significant effects. Among the AM technologies tested, all have produced either comparable or truer 3D-printed dies than the Type IV dental stone dies, and the CLIP and SLA 3D-printers produced the best outcomes. 3D-printed dies showed significant distortion after 1-month and 3-months storage, especially under light exposure storage conditions. These findings may negate the clinical need to preserve 3D-printed dies, and digital data should be preserved instead.

Long-term results from the multicentric European randomized phase 3 trial CWS/RMS-96 for localized high-risk soft tissue sarcoma in children, adolescents, and young adults.

BACKGROUND: CWS/RMS-96 was an international multicenter trial with randomization between two therapy arms of the standard four-drug therapy (vincristine, ifosfamide, adriamycin, dactinomycin [VAIA]) versus an intensified six-drug regimen (carboplatin, epirubicin, vincristine, dactinomycin, ifosfamide, and etoposide [CEVAIE]) for high-risk rhabdomyosarcoma (RMS), extraskeletal Ewing sarcoma (EES), and undifferentiated sarcoma (UDS) in children, adolescents, and young adults aiming to improve their survival. Intensified chemotherapy with CEVAIE did not improve outcome. METHODS: Patients younger than 21 years with a previously untreated localized HR-RMS, EES, and UDS were enrolled from Cooperative Weichteilsarkom Studiengruppe (CWS) centers in Germany, Austria, Poland, Switzerland, and from Italian Soft Tissue Sarcoma Committee (STSC) centers. Randomization (1:1) to receive either 9 × 21 days cycles of VAIA or CEVAIE was performed separately in CWS and STSC. Hyperfractionated accelerated radiotherapy (32-44.8 Gy) was added at week 9-12 according to histology and response to chemotherapy. A secondary microscopically complete nonmutilating resection was performed if possible. Primary endpoints were response to chemotherapy, event-free (EFS) and overall survival (OS). RESULTS: Five hundred fifty-seven patients (HR-RMS: n = 416, EES and UDS: n = 141) underwent randomization: VAIA (n = 273) or CEVAIE (n = 284). Radiotherapy was given to 70% of patients in both groups. A secondary resection was performed in 47% and 48% patients, respectively. The 5-year EFS and OS for the VAIA and CEVAIE treatment arms were 59.8% and 60.8% (p = .89), and 74.2% and 68.3% (p = .16), respectively. No differences in response, toxicity, or second malignancies emerged in the two groups. CONCLUSION: The use of an intensified regimen failed to show a significant improvement in tumor response and outcome of patients with localized HR-RMS, EES, and UDS.

Italian translation and validation of fatigue symptoms and impacts questionnaire in relapsing multiple sclerosis (FSIQ-RMS).

BACKGROUND: Fatigue is a disabling symptom of multiple sclerosis (MS) and impacts on daily life. The Fatigue Symptoms and Impacts Questionnaire-Relapsing Multiple Sclerosis (FSIQ-RMS) is a new 20-item tool that investigates the aspects of fatigue in MS. It concerns motor, cognitive, social, and emotional aspects of fatigue. We aim at validating the Italian version of FSIQ-RMS in an Italian population of MS patients and demonstrate its reliability and construct validity. METHODS: We included patients with diagnosis of MS, age between 18 and 70 years with ability to participate in a 90-min interview. Each patient completed the Italian version of FSIQ-RMS and Fatigue Severity Scale (FSS) at the same time. Construct validity was explored by the exploratory factor analysis; reliability was assessed with Cronbach's alpha; and test-retest stability was examined through intraclass correlation coefficient (ICC). Concurrent validity was calculated using Pearson's correlation. RESULTS: We enrolled 171 patients (126 female and 45 male), 83% with relapsing MS (RMS), and 17% with secondary progressive MS (SPMS). Italian FSIQ-RMS showed a Cronbach's alpha of 0.92; ICC was 0.96. Pearson's correlation coefficient between FSIQ-RMS and FSS total score was statistically significant (p < 0.01); FSIQ-RMS inversely correlated also with BMI and positively with EDSS. CONCLUSION: The Italian version of FSIQ-RMS has excellent psychometric properties and can be used in research and clinical setting to evaluate physical, cognitive, and social fatigue in both RMS and SPMS.

Combined mTORC1/mTORC2 inhibition blocks growth and induces catastrophic macropinocytosis in cancer cells.

The mammalian target of rapamycin (mTOR) pathway, which plays a critical role in regulating cellular growth and metabolism, is aberrantly regulated in the pathogenesis of a variety of neoplasms. Here we demonstrate that dual mTORC1/mTORC2 inhibitors OSI-027 and PP242 cause catastrophic macropinocytosis in rhabdomyosarcoma (RMS) cells and cancers of the skin, breast, lung, and cervix, whereas the effects are much less pronounced in immortalized human keratinocytes. Using RMS as a model, we characterize in detail the mechanism of macropinocytosis induction. Macropinosomes are distinct from endocytic vesicles and autophagosomes in that they are single-membrane bound vacuoles formed by projection, ruffling, and contraction of plasma membranes. They are positive for EEA-1 and LAMP-1 and contain watery fluid but not organelles. The vacuoles then merge and rupture, killing the cells. We confirmed the inhibition of mTORC1/mTORC2 as the underpinning mechanism for macropinocytosis. Exposure to rapamycin, an mTORC1 inhibitor, or mTORC2 knockdown alone had little or reduced effect relative to the combination. We further demonstrate that macropinocytosis depends on MKK4 activated by elevated reactive oxygen species. In a murine xenograft model, OSI-027 reduced RMS tumor growth. Molecular characterization of the residual tumors was consistent with the induction of macropinocytosis. Furthermore, relative to the control xenograft tumors, the residual tumors manifested reduced expression of cell proliferation markers and proteins that drive the epithelial mesenchymal transition. These data indicate a role of mTORC2 in regulating tumor growth by macropinocytosis and suggest that dual inhibitors could help block refractory or recurrent RMS and perhaps other neoplasms and other cancer as well.