Role of catecholamine synthases in the maintenance of cancer stem-like cells in malignant peripheral nerve sheath tumors.

Malignant peripheral nerve sheath tumors (MPNSTs) are malignant tumors that are derived from Schwann cell lineage around peripheral nerves. As in many other cancer types, cancer stem cells (CSCs) have been identified in MPNSTs, and they are considered the cause of treatment resistance, recurrence, and metastasis. As an element defining the cancer stemness of MPNSTs, we previously reported a molecular mechanism by which exogenous adrenaline activates a core cancer stemness factor, YAP/TAZ, through ß2 adrenoceptor (ADRB2). In this study, we found that MPNST cells express catecholamine synthases and that these enzymes are essential for maintaining cancer stemness, such as the ability to self-renew and maintain an undifferentiated state. Through gene knockdown and inhibition of these enzymes, we confirmed that catecholamines are indeed synthesized in MPNST cells. The results confirmed that catecholamine synthase knockdown in MPNST cells reduces the activity of YAP/TAZ. These data suggest that a mechanism of YAP/TAZ activation by de novo synthesized adrenaline, as well as exogenous adrenaline, may exist in the maintenance of cancer stemness of MPNST cells. This mechanism not only helps to understand the pathology of MPNST, but could also contribute to the development of therapeutic strategies for MPNST.

PRRX1-TOP2A interaction is a malignancy-promoting factor in human malignant peripheral nerve sheath tumours.

BACKGROUND: Paired related-homeobox 1 (PRRX1) is a transcription factor in the regulation of developmental morphogenetic processes. There is growing evidence that PRRX1 is highly expressed in certain cancers and is critically involved in human survival prognosis. However, the molecular mechanism of PRRX1 in cancer malignancy remains to be elucidated. METHODS: PRRX1 expression in human Malignant peripheral nerve sheath tumours (MPNSTs) samples was detected immunohistochemically to evaluate survival prognosis. MPNST models with PRRX1 gene knockdown or overexpression were constructed in vitro and the phenotype of MPNST cells was evaluated. Bioinformatics analysis combined with co-immunoprecipitation, mass spectrometry, RNA-seq and structural prediction were used to identify proteins interacting with PRRX1. RESULTS: High expression of PRRX1 was associated with a poor prognosis for MPNST. PRRX1 knockdown suppressed the tumorigenic potential. PRRX1 overexpressed in MPNSTs directly interacts with topoisomerase 2 A (TOP2A) to cooperatively promote epithelial-mesenchymal transition and increase expression of tumour malignancy-related gene sets including mTORC1, KRAS and SRC signalling pathways. Etoposide, a TOP2A inhibitor used in the treatment of MPNST, may exhibit one of its anticancer effects by inhibiting the PRRX1-TOP2A interaction. CONCLUSION: Targeting the PRRX1-TOP2A interaction in malignant tumours with high PRRX1 expression might provide a novel tumour-selective therapeutic strategy.

Changes in Spinal Instability After Conventional Radiotherapy for Painful Vertebral Bone Metastases.

OBJECTIVE: Precise assessment of spinal instability is critical before and after radiotherapy (RT) for evaluating the effectiveness of RT. Therefore, we retrospectively evaluated the efficacy of RT in spinal instability over a period of 6 months after RT, utilizing the spinal instability neoplastic score (SINS) in patients with painful spinal metastasis. We retrospectively evaluated 108 patients who received RT for painful vertebral metastasis in our institution. Mechanical pain at metastatic vertebrae, radiological responses of irradiated vertebrae, and spinal instability were assessed. Follow-up assessments were done at the start of and at intervals of 1, 2, 3, 4, and 6 months after RT, with the pain disappearing in 67%, 85%, 93%, 97%, and 100% of the patients, respectively. The median SINS were 8, 6, 6, 5, 5, and 4 at the beginning and after 1, 2, 3, 4, and 6 months of RT, respectively. Multivariate analysis revealed that posterolateral involvement of spinal elements (PLISE) was the only risk factor for continuous potentially unstable/unstable spine at 1 month. In conclusion, there was improvement of pain, and recalcification results in regaining spinal stability over time after RT although vertebral body collapse and malalignment occur in some irradiated vertebrae. Clinicians should pay attention to PLISE in predicting continuous potentially unstable/unstable spine.

Circularly polarised luminescence from intramolecular excimer emission of bis-1,8-naphthalimide derivatives.

Chiral excimers exhibit unique photophysical behaviour. However, further molecular design is required along with systematic studies on the effect of spacer groups and solvent polarity. In this study, we prepared four circularly polarised luminescence (CPL)-active molecules that exhibit intramolecular excimer emission. Bis-1,8-naphthalimide (bNI) derivatives D-LybNI, L-LybNI, D-LyMebNI, and L-LyMebNI were prepared with chiral backbones and alkyl linkages between the NI rings with chain lengths of five carbon atoms, suitable for excimer fluorescence. The fluorescence properties were investigated experimentally and theoretically using density functional theory. The molecules exhibited intramolecular excimer fluorescence in polar organic solvents. Mirror-image circular dichroism and CPL spectra were obtained for the D and L forms. D- and L-LyMebNI exhibited relatively large luminescence dissymmetry factors (|glum|) in acetonitrile of 1.9 × 10-3 and 1.6 × 10-3, respectively. Thus, this study demonstrates chiral bNI derivatives with simple synthesis procedures that emit intramolecular excimer fluorescence and have effective CPL properties. These molecules are promising for developing organic molecular systems with bright, highly polarised emission.

Macropinoscope: Real-Time Simultaneous Tracking of pH and Cathepsin B Activity in Individual Macropinosomes.

A fluorescent sensor that allows simultaneous analysis of environmental factors in a limited cellular space is useful for understanding precise molecular interactions in live cells and their biological responses. Macropinocytosis is a ubiquitous endocytic pathway for massive uptake of extracellular fluids, resulting in the formation of macropinosomes. Although macropinocytosis may impact intracellular delivery and cancer proliferation, information on the intracellular behaviors of macropinosomes is limited. Here, we aimed to develop a macropinoscope, a sensor that simultaneously detects pH and cathepsin B activity in individual macropinosomes. A macropinosome-specific marker, dextran (70 kDa), was employed as a platform, onto which fluorescein, Oregon Green, and tetramethylrhodamine were loaded for ratiometric pH sensing and imaging. A cathepsin-B-cleavable peptide sequence bearing sulfo-Cy5 and the quencher BHQ-3 was also mounted; cleavage of the sequence was detected as an increase in sulfo-Cy5 fluorescence. A steep decrease in pH was observed 5-10 min after macropinosome formation, which was accompanied by an immediate increase in cathepsin B activity. Our design concept will lead to the development of other macropinoscopes for the simultaneous detection of other parameters in individual macropinosomes.

An Artificial Liposome Compartment with Size Exclusion Molecular Transport.

The cellular compartment plays an essential role in organizing the complex and diverse biochemical reactions within the cell. By mimicking the function of such cellular compartments, the challenge of constructing artificial compartments has been taken up to develop new biochemical tools for efficient material production and diagnostics. The important features required for the artificial compartment are that it isolates the interior from the external environment and is further functionalized to control the transport of target chemicals to regulate the interior concentration of both substrate and reaction products. In this study, an artificial compartment with size-selective molecular transport function was constructed by using a DNA origami-guided liposome prepared by modifying the method reported by Perrault et al. This completely isolates the liposome interior, including the DNA origami skeleton, from the external environment and allows the assembly of a defined number of molecules of interest inside and/or outside the compartment. By incorporating a bacterial membrane protein, OmpF, into the liposome, the resulting artificial compartment was shown to transport only the molecule of interest with a molecular weight below 600 Da from the external environment into the interior of the compartment.

Advances in treatment of alveolar soft part sarcoma: an updated review.

Alveolar soft part sarcoma is a rare neoplasm of uncertain histogenesis that belongs to a newly defined category of ultra-rare sarcomas. The neoplasm is characterized by a specific chromosomal translocation, der (17) t(X; 17)(p11.2;q25), that results in ASPSCR1-TFE3 gene fusion. The natural history of alveolar soft part sarcoma describes indolent behaviour with slow progression in deep soft tissues of the extremities, trunk and head/neck in adolescents and young adults. A high rate of detection of distant metastasis at presentation has been reported, and the most common metastatic sites in decreasing order of frequency are the lung, bone and brain. Complete surgical resection remains the standard treatment strategy, whereas radiotherapy is indicated for patients with inadequate surgical margins or unresectable tumours. Although alveolar soft part sarcoma is refractory to conventional doxorubicin-based chemotherapy, monotherapy or combination therapy using tyrosine kinase inhibitors and immune checkpoint inhibitors have provided antitumor activity and emerged as new treatment strategies. This article provides an overview of the current understanding of this ultra-rare sarcoma and recent advancements in treatments according to the clinical stage of alveolar soft part sarcoma.

Soft-tissue sarcoma in adolescents and young adults.

Soft-tissue sarcoma is a rare cancer that accounts for approximately 1% of all malignant tumors. Although they occur in various age groups, soft-tissue sarcomas account for 8% of all malignant tumors developing in adolescents and young adults, suggesting that they are not rare in this age group. This study aimed to evaluate the clinical and pathological characteristics of soft-tissue sarcoma in adolescents and young adults. According to the Bone and Soft-Tissue Tumor Registry in Japan, myxoid liposarcoma is the most common type of soft-tissue sarcoma found in adolescents and young adults; alveolar soft part sarcoma, extraskeletal Ewing sarcoma, epithelioid sarcoma, clear cell sarcoma and synovial sarcoma occur predominantly in this age group among soft-tissue sarcomas. The analysis based on this registry demonstrated that age was not a prognostic factor for poor survival of soft-tissue sarcoma, although the prognosis in adolescents and young adults was better than that in older patients in the US and Scandinavia. Adolescent and young adult patients with soft-tissue sarcoma have age-specific problems, and a multidisciplinary approach to physical, psychological, and social issues is necessary to improve the management of these young patients both during and after treatment.

Stabilization and structural changes of 2D DNA origami by enzymatic ligation.

The low thermal stability of DNA nanostructures is the major drawback in their practical applications. Most of the DNA nanotubes/tiles and the DNA origami structures melt below 60°C due to the presence of discontinuities in the phosphate backbone (i.e., nicks) of the staple strands. In molecular biology, enzymatic ligation is commonly used to seal the nicks in the duplex DNA. However, in DNA nanotechnology, the ligation procedures are neither optimized for the DNA origami nor routinely applied to link the nicks in it. Here, we report a detailed analysis and optimization of the conditions for the enzymatic ligation of the staple strands in four types of 2D square lattice DNA origami. Our results indicated that the ligation takes overnight, efficient at 37°C rather than the usual 16°C or room temperature, and typically requires much higher concentration of T4 DNA ligase. Under the optimized conditions, up to 10 staples ligation with a maximum ligation efficiency of 55% was achieved. Also, the ligation is found to increase the thermal stability of the origami as low as 5°C to as high as 20°C, depending on the structure. Further, our studies indicated that the ligation of the staple strands influences the globular structure/planarity of the DNA origami, and the origami is more compact when the staples are ligated. The globular structure of the native and ligated origami was also found to be altered dynamically and progressively upon ethidium bromide intercalation in a concentration-dependent manner.

Navicular Bone Fracture after Radiofrequency Ablation in a Patient with Osteoid Osteoma.

Osteoid osteoma (OO) is a benign bone tumor that presents with nocturnal pain. Computed tomography (CT)- guided radiofrequency ablation (RFA) has been widely performed for OO, and major adverse events post-RFA are rare. We report a case of OO in the left navicular bone of a 15-year-old male. He underwent RFA for OO, and the pain improved temporarily. At the 1-month follow-up, the patient complained of left foot pain, and a CT examination revealed a fracture of the ablated navicular bone. Fractures are rare but must be taken into account after bone RFA.

Comparison between Cases of Total Hip Arthroplasty Followed by Colonna Capsular Arthroplasty and Lorenz Cast Reduction in Patients with Developmental Dysplasia of the Hip.

Most patients with developmental dysplasia of the hip (DDH) now receive closed-reduction treatment within 6 months after birth. The long-term outcomes of patients with late-detection DDH have remained unclear. We reviewed the clinical records of 18 patients who underwent Colonna capsular arthroplasty (n=8) or closed reduction (n=10) for developmental dysplasia of the hip as infants or young children and underwent total hip arthroplasty approximately in midlife. Both the Colonna capsular arthroplasty and closed reduction groups achieved good clinical results after total hip arthroplasty. However, the operating time was longer and the improvements of hip range of motion and clinical score were significantly worse in the Colonna capsular arthroplasty group than in the closed reduction group.

A Boy Safely Treated with Tyrosine Kinase Inhibitors for Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia with Osteolysis.

A three-year-old boy with Philadelphia chromosome-positive B-cell precursor acute lymphoblastic leukemia (Ph+ALL) presented with an osteolytic lesion in his right upper arm. Tyrosine kinase inhibitors (TKIs) such as imatinib and dasatinib are an essential component throughout the course of treatment for Ph+ALL. However, TKIs are reported to affect the bone metabolism. In the treatment course of the current patient, the osteolytic lesion quickly improved despite the continuous use of TKIs, even during the concomitant use of corticosteroids. This suggests that TKIs can be safely given with concomitant corticosteroids to children with Ph+ALL, even when osteolytic lesions are present.

Controlled Assembly of Fluorophores inside a Nanoliposome.

Cellular compartmentalization plays an essential role in organizing the complex and multiple biochemical reactions in the cell. An artificial compartment would provide powerful strategies to develop new biochemical tools for material production and diagnosis, but it is still a great challenge to synthesize the compartments that encapsulate materials of interest while controlling their accurate locations, numbers, and stoichiometry. In this study, we evaluated chemical characteristics of a liposome-encapsulated compartment, which has great potential to locate various materials of interest with precise control of their locations and numbers in the compartment. A nanoliposome was constructed inside a ring-shaped DNA origami skeleton according to the method of Yang et al., and further equipped with a double-stranded DNA platform to assemble molecules of interest in the nanoliposome. Upon formation of the nanoliposome, a pH-sensitive fluorophore on the bridged platform showed little or no response to the pH change of the outer buffer, ensuring that the molecules assembled on the platform are effectively shielded from the outer environment. The ring-shaped DNA skeleton equipped with a double-stranded DNA platform allows spatial assembly of several functional molecules inside the nanoliposome to isolate them from the outer environment.

FRET-Based Cathepsin Probes for Simultaneous Detection of Cathepsin B and D Activities.

Fluorescent cathepsin probes were prepared by modification of peptidic substrates for cathepsin B (CTSB) and cathepsin D (CTSD) with FRET pairs. Fluorophores with distinguishable emission characteristics were applied to CTSB and CTSD probes with their appropriate quenchers to simultaneously monitor the activity of CTSB and/or CTSD. Conjugation of both the CTSB and CTSD probes with short single-stranded DNA drastically increased their reactivity to cathepsins over the parent probes possibly by improving their solubility. The activity of CTSB and CTSD were simultaneously detected by using these orthogonal FRET-based cathepsin probes.

Topologically-Interlocked Minicircles as Probes of DNA Topology and DNA-Protein Interactions.

DNA minicircles exist in biological contexts, such as kinetoplast DNA, and are promising components for creating functional nanodevices. They have been used to mimic the topological features of nucleosomal DNA and to probe DNA-protein interactions such as HIV-1 and PFV integrases, and DNA gyrase. Here, we synthesized the topologically-interlocked minicircle rotaxane and catenane inside a frame-shaped DNA origami. These minicircles are 183 bp in length, constitute six individual single-stranded DNAs that are ligated to realize duplex interlocking, and adopt temporary base pairing of single strands for interlocking. To probe the DNA-protein interactions, restriction reactions were carried out on DNAs with different topologies such as free linear duplex or duplex constrained inside origami and free or topologically-interlocked minicircles. Except the free linear duplex, all tested structures were resistant to restriction digestion, indicating that the topological features of DNA, such as flexibility, curvature, and groove orientation, play a major role in DNA-protein interactions.

Topologically-Interlocked Minicircles as Probes of DNA Topology and DNA-Protein Interactions.

Invited for the cover of this issue are Prof. Takashi Morii and co-workers at Kyoto University and Ewha Womans University. The cover image depicts the graphical design and atomic force microscopic (AFM) images of the synthesized topologically-interlocked DNA catenane and rotaxanes inside a frame-shaped DNA origami. Read the full text of the article at 10.1002/chem.202200108.

Alveolar soft part sarcoma: progress toward improvement in survival? A population-based study.

BACKGROUND: Alveolar soft part sarcoma (ASPS) is a rare histological subtype of soft-tissue sarcoma, which remains refractory to conventional cytotoxic chemotherapy. We aimed to characterize ASPS and investigate whether the oncological outcome has improved over the past decade. METHODS: One hundred and twenty patients with newly diagnosed ASPS from 2006 to 2017, identified from the Bone and Soft-Tissue Tumor Registry in Japan, were analyzed retrospectively. RESULTS: The study cohort comprised 34 (28%) patients with localized ASPS and 86 (72%) with metastatic disease at presentation. The 5-year disease-specific survival (DSS) was 68% for all patients and 86% and 62% for localized and metastatic disease, respectively (p = 0.019). Metastasis at presentation was the only adverse prognostic factor for DSS (hazard ratio [HR]: 7.65; p = 0.048). Patients who were > 25 years (80%; p = 0.023), had deep-seated tumors (75%; p = 0.002), and tumors > 5 cm (5-10 cm, 81%; > 10 cm, 81%; p < 0.001) were more likely to have metastases at presentation. In patients with localized ASPS, adjuvant chemotherapy or radiotherapy did not affect survival, and 13 patients (45%) developed distant metastases in the lung (n = 12, 92%) and brain (n = 2, 15%). In patients with metastatic ASPS (lung, n = 85 [99%]; bone, n = 12 [14%]; and brain n = 9 [11%]), surgery for the primary or metastatic site did not affect survival. Prolonged survival was seen in patients who received pazopanib treatment (p = 0.045), but not in those who received doxorubicin-based cytotoxic chemotherapy. Overall, improved DSS for metastatic ASPS has been observed since 2012 (5-year DSS, from 58 to 65%) when pazopanib was approved for advanced diseases, although without a statistically significant difference (p = 0.117). CONCLUSION: The national study confirmed a unique feature of ASPS with frequent metastasis to the lung and brain but an indolent clinical course. An overall trend toward prolonged survival after the introduction of targeted therapy encourages continuous efforts to develop novel therapeutic options for this therapeutically resistant soft-tissue sarcoma.

Short-term clinical outcomes of Kyocera Modular Limb Salvage System designed cementless stems for the endoprosthetic reconstruction of lower extremities: a Japanese Musculoskeletal Oncology Group multi-institutional study.

BACKGROUND: The high rate of aseptic loosening of cemented stems has led to their frequent use in endoprosthetic reconstruction. However, problems, such as stem breakage and stress shielding at the insertion site, remain. The Japanese Musculoskeletal Oncology Group (JMOG) has developed Kyocera Modular Limb Salvage System (KMLS) cementless stems with a unique tapered press-fit and short fixation design. This study aimed to clarify the short-term postoperative outcomes of this prosthesis and validate the stem design. METHODS: One hundred cases of KMLS cementless stems (51 male patients; median age, 49 years; mean follow-up period, 35 months), with a minimum follow-up of 2 years, for the proximal femur (PF), distal femur (DF), and proximal tibia were prospectively registered for use. Prosthesis survival, complication rates, postoperative functional, and radiographical evaluation were analyzed. Complications or failures after insertion of the KMLS endoprostheses were classified into five types and functional results were analyzed according to the MSTS scoring system at postoperative 1 year. The diaphyseal interface and anchorage were graded by the ISOLS system at postoperative 2 years. RESULTS: The overall prosthesis survival rates at 2 and 4 years were 88.2 and 79.6%, respectively. The prosthesis-specific survival rate excluding infection and tumor recurrence was 90.2 and 87.9%, respectively. Younger age (p = 0.045) and primary tumor (p = 0.057) were associated with poor prognosis of prosthesis-specific survival excluding infection and tumor recurrence. Complications were observed in 31 patients, 13 patients underwent revision surgery. The mean MSTS functional score at 1 year postoperatively was 68%. Early implant loosening was significantly more common in the DF (p = 0.006) and PF/DF straight stem (p = 0.038). The ISOLS radiographic evaluation at 2 years after surgery revealed good bone remodeling and anchorage in most cases (bone remodeling: 90% / excellent and good, anchorage: 97% / excellent and good). CONCLUSIONS: Tumor endoprosthesis long-term fixation to the diaphysis of the lower extremity remains challenging. The KMLS cementless stem with a unique tapered press fit design showed good short-term results in maintaining bone stock. To prevent early loosening, a curved stem should be used in PF and DF, but long-term follow-up is necessary.

Ultrasound-dependent RNAi using TatU1A-rose bengal conjugate.

Tat-U1A-rose bengal conjugate (TatU1A-RB) was prepared as an ultrasound-sensitive RNA carrier molecule. This molecule consists of Tat cell-penetrating peptide, U1A RNA-binding protein, and rose bengal as a sonosensitizer. We demonstrated that TatU1A-RB delivered RNA via the endocytosis pathway, which was followed by ultrasound-dependent endosomal escape and cytosolic dispersion of the RNA. A short hairpin RNA (shRNA) delivered by TatU1A-RB mediated RNA interference (RNAi) ultrasound-dependently. Even by ultrasound irradiation through blood cells, RNAi could be induced with TatU1A-RB and the shRNA. This ultrasound-dependent cytosolic RNA delivery method will serve as the basis for a new approach to nucleic acid therapeutics.

Grade of Subchondral Insufficiency Fracture of the Knee and the Presence of a Posterior Shiny-Corner Lesion are Correlated with Duration of Medial Meniscus Posterior Root Tear in Women.

Bone marrow edema (BME) after meniscus injury and risk factors for subchondral insufficiency fracture of the knee (SIFK) have been reported. However, their association with medial meniscus posterior root tear (MMPRT) remains unknown. We investigated the association of BME volume (BME-V), posterior shinycorner lesion (PSCL), and SIFK with MMPRT to examine the correlations between BME-V and medial meniscus extrusion (MME), PSCL and duration from injury to the time of magnetic resonance imaging (duration), and SIFK and duration. Twenty-nine patients who underwent surgery for MMPRT were included (mean age, 59.2; range, 39-84). The presence of PSCL, femoral BME-V (cm3), and SIFK grade (1-4) were evaluated. Preoperative factors, such as MME (mm) and duration (weeks), were investigated using multivariate linear/ logistic regression analyses. Multivariate linear regression analysis revealed duration as a significant factor for high-grade SIFK (p<0.01). Multivariate logistic regression analysis revealed duration as a significant factor for the presence of PSCL (odds ratio=0.94, p<0.05). A long duration of MMPRT leads to severe MME and highgrade SIFK (3 and 4), often resulting in knee arthroplasty. Early diagnosis of MMPRT and pullout repair can prevent severe MME and high-grade SIFK.