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Abstract Background/Aim For patients treated with osimertinib as first-line therapy, there have been no studies comparing both progression-free survival (PFS) and overall survival (OS) according to performance status (PS). Furthermore, no studies have examined differences in baseline genetic abnormalities between patients with poor and good PS. Therefore, we aimed to investigate differences in baseline genetic abnormalities and treatment effects between patients with poor and good PS who received osimertinib as the primary treatment. Patients and methods This is a secondary analysis of the ELUCIDATOR study, which is a multi-center prospective observational study in Japan that assessed mechanisms underlying resistance to osimertinib as first-line treatment for advanced non-small cell lung cancer with epidermal growth factor receptor mutations. Results There were 153 and 25 patients in the good and poor PS groups, respectively. Multivariate analysis revealed no significant between-group differences in PFS (hazards ratio [HR]: 0.98, 95% confidence interval [CI]: 0.52–1.72, p = 0.946). Multivariate analysis of OS revealed that poor PS was a poor prognostic factor (HR: 2.67, 95% CI: 1.43–4.73, p = 0.003). Regarding baseline genetic abnormalities, there was a significant increase in APC-positive cases (20.0% vs. 2.2%, p = 0.009) and a trend toward more CTNNB1-positive cases in the poor PS group than in the good PS group (14.3% vs. 2.9%, p = 0.062). Conclusion There was no between-group difference in PFS, although OS was significantly inferior in the poor PS group. Additionally, there was a significant increase in APC-positive cases and a trend toward more CTNNB1-positive cases in the poor PS group.

Summary: Point mutations in KIF22 have been linked to spondyloepimetaphyseal dysplasia with joint laxity, type 2 (SEMDJL2). Skeletal features of SEMDJL2 include short stature and joint laxity. Mechanisms underlying these limb abnormalities are unknown. Here in this manuscript, we have investigated the function of KIF22 in chondrocytes. Quantitative PCR and immunostaining revealed that Kif22 was highly expressed in proliferating-zone growth-plate chondrocytes. Kif22 knockdown resulted in defective mitotic spindle formation and reduced cell proliferation. Forced expression of SEMDJL-associated mutant Kif22 constructs likewise induced abnormal mitotic spindle morphology and reduced proliferation. Mice expressing a KIF22 truncation mutant had shorter growth plates and shorter tibial bones compared to wild-type mice. These results suggest that KIF22 regulates mitotic spindle formation in proliferating chondrocytes thereby linking the stunted longitudinal bone growth observed in SEMDJL2 to failures of chondrocyte division.

Objective The application of the da Vinci Surgical System in neurosurgery is limited due to technical difficulties requiring precise maneuvers and small instruments. This study details the advantages and disadvantages of robotics in neurosurgery and the reachable range of the transoral approach to lesions of the skull base and upper cervical spine. Methods In a cadaver study, the da Vinci Xi robot, lacking haptic feedback, was utilized for sagittal and coronal approaches on 5 heads, facilitating dura suturing in 3, with a 30°-angled drill for bone removal. Results Perfect exposure of all the nasopharyngeal sites, clivus, sellar, and choana, including the bilateral eustachian tubes, was achieved without any external incisions using this palatal split approach of transoral robotic surgery. The time required to perform a single stitch, knot, and complete single suture in robotic suturing of deep-seated were significantly less compared to manual suturing via the endonasal approach. Conclusion This is the first report to show the feasibility of suturing the dural defect in deep-seated lesions transorally and revealed that the limit of reach in the coronal plane via a transoral approach with incision of the soft palate is the foramen ovale. This preclinical investigation also showed that the transoral robotic approach is feasible for lesions extending from the sellar to the C2 in the sagittal plane. Refinement of robotic instruments for specific anatomic sites and future neurosurgical studies are needed to further demonstrate the feasibility and effectiveness of this system in treating benign and malignant skull base lesions.

Abstract Introduction Combined treatment using anti‐programmed death‐ligand 1 antibody (anti‐PD‐L1) and platinum‐etoposide is the current standard first‐line treatment for patients with extensive‐stage (ES) small cell lung cancer (SCLC). However, the best treatment for relapsed ES‐SCLC after the first‐line treatment remains unclear. There are some approved chemotherapeutic agents that can be used against ES‐SCLC, and treatment with irinotecan is well established as both a monotherapy and a combined therapy, in combination with platinum. Therefore, we conduct a phase II study with irinotecan in the second‐ or later‐line setting for patients with ES‐SCLC who have been previously treated with combined treatment. Methods Our study will enroll total 30 patients who are diagnosed with ES‐SCLC and have experienced disease progression after the combined treatment. Patients will receive irinotecan on days 1, 8, and 15, which will be repeated every 4 weeks. Doses of irinotecan (100/80/60 mg/m2) will be determined according to the type of UGT1A1 gene polymorphism, and the treatment will be discontinued following disease progression, intolerance, withdrawal of patient consent, and based on the investigator's decision. The primary endpoint of the study is the response rate, and the secondary endpoints are overall survival, progression‐free survival, and safety. Discussion Since the present first‐line treatment has been changed to the combined treatment, the second‐ or later‐line treatment should be re‐evaluated for patients with relapsed SCLC. Irinotecan is a major chemotherapeutic agent used for SCLC. This study demonstrates and re‐evaluates the clinical benefits of irinotecan after combined treatment with anti‐PD‐L1 and platinum‐etoposide for patients with ES‐SCLC. Registration details This study was registered in the Japan Registry of Clinical Trials (no. jRCT s071210090) on November 4, 2021.

Abstract Although recent molecular analyses revealed that sporadic meningiomas have various genetic, epigenetic, and transcriptomic profiles, meningioma in patients with neurofibromatosis type 2 (NF2) have not been fully elucidated. This study investigated meningiomas' clinical, histological, and molecular characteristics in NF2 patients. A long-term retrospective follow-up (13.5 ± 5.5 years) study involving total 159 meningiomas in 37 patients with NF2 was performed. Their characteristics were assessed using immunohistochemistry (IHC), bulk-RNA sequencing, and copy number analysis. All variables of meningiomas in patients with NF2 were compared with those in 189 sporadic NF2-altered meningiomas in 189 patients. Most meningiomas in NF2 patients were stable, and the mean annual growth rate was 1.0 ± 1.8 cm3/year. Twenty-eight meningiomas (17.6%) in 25 patients (43.1%) were resected during the follow-up period. WHO grade I meningiomas in patients with NF2 were more frequent than in sporadic NF2-altered meningiomas (92.9% vs. 80.9%). Transcriptomic analysis for patients with NF2/sporadic NF2-altered WHO grade I meningiomas (n = 14 vs. 15, respectively) showed that tumours in NF2 patients still had a higher immune response and immune cell infiltration than sporadic NF2-altered meningiomas. Furthermore, RNA-seq/IHC-derived immunophenotyping corroborated this enhanced immune response by identifying myeloid cell infiltration, particularly in macrophages. Clinical, histological, and transcriptomic analyses of meningiomas in patients with NF2 demonstrated that meningiomas in NF2 patients showed less aggressive behaviour than sporadic NF2-altered meningiomas and elicited a marked immune response by identifying myeloid cell infiltration, particularly of macrophages.

Abstract Background Dacomitinib significantly improves progression‐free survival and overall survival (OS) compared with gefitinib in patients with non–small‐cell lung cancer (NSCLC) harboring epidermal growth factor receptor (EGFR)‐activating mutations. However, dacomitinib often causes skin toxicities, resulting in treatment discontinuation. We aimed to evaluate a prophylactic strategy for skin toxicity induced by dacomitinib. Methods We performed a single‐arm, prospective, open‐label, multi‐institutional phase II trial for comprehensive skin toxicity prophylaxis. Patients with NSCLC harboring EGFR‐activating mutations were enrolled and received dacomitinib with comprehensive prophylaxis. The primary endpoint was the incidence of skin toxicity (Grade ≥2) in the initial 8 weeks. Results In total, 41 Japanese patients participated between May 2019 and April 2021 from 14 institutions (median age 70 years; range: 32–83 years), 20 were male, and 36 had a performance status of 0–1. Nineteen patients had exon 19 deletions and L858R mutation. More than 90% of patients were perfectly compliant with prophylactic minocycline administration. Skin toxicities (Grade ≥2) occurred in 43.9% of patients (90% confidence interval [CI], 31.2%–56.7%). The most frequent skin toxicity was acneiform rash in 11 patients (26.8%), followed by paronychia in five patients (12.2%). Due to skin toxicities, eight patients (19.5%) received reduced doses of dacomitinib. The median progression‐free survival was 6.8 months (95% CI, 4.0–8.6 months) and median OS was 21.6 months (95% CI, 17.0 months–not reached). Conclusion Although the prophylactic strategy was ineffective, the adherence to prophylactic medication was quite good. Patient education regarding prophylaxis is important and can lead to improved treatment continuity.

Abstract Vitamin E is classified into tocopherol (Toc) and tocotrienol (T3) based on its side chains. T3 generally has higher cellular uptake than Toc, though the responsible mechanism remains unclear. To elucidate this mechanism, we hypothesized and investigated whether serum albumin is a factor that induces such a difference in the cellular uptake of Toc and T3. Adding bovine serum albumin (BSA) to serum-depleted media increased the cellular uptake of T3 and decreased that of Toc, with varying degrees among α-, β-, γ-, and δ-analogs. Such enhanced uptake of α-T3 was not observed when cells were incubated under low temperature (the uptake of α-Toc was also reduced), suggesting that Toc and T3 bind to albumin to form a complex that results in differential cellular uptake of vitamin E. Fluorescence quenching study confirmed that vitamin E certainly bound to BSA, and that T3 showed a higher affinity than Toc. Molecular docking further indicated that the differential binding energy of Toc or T3 to BSA is due to the Van der Waals interactions via their side chain. Overall, these results suggested that the affinity of Toc and T3 to albumin differs due to their side chains, causing the difference in their albumin-mediated cellular uptake. Our results give a better mechanistic insight into the physiological action of vitamin E.

BackgroundDespite accumulating research on the molecular characteristics of meningiomas, no definitive molecularly targeted therapy for these tumors has been established to date. Molecular mechanisms underlying meningioma progression also remain unclear. Comprehensive genetic testing approaches can reveal actionable gene aberrations in meningiomas. However, there is still limited information on whether profiling the molecular status of subsequent recurrent meningiomas could influence the choice of molecular-targeted therapies.Case presentationWe report a case of meningioma with malignant progression and multiple recurrences. We performed matched tumor pair analysis using the Todai OncoPanel to investigate the possibility of additional standard treatments. The loss of several chromosomal regions, including NF2 and CDKN2A, which is associated with aggressive meningiomas, was considered a significant driver event for malignant progression. Using additional matched tumor pair analysis, mutations in TRAF7, ARID1A, and ERBB3 were identified as subclonal driver events at the time of recurrence. No genetic aberrations were found for which evidence-based targeted therapy was applicable. We also reviewed previous reports of molecular therapies in meningioma to discuss issues with the current molecular testing approach.ConclusionGene panel testing platforms such as the Todai OncoPanel represent a powerful approach to elucidate actionable genetic alterations in various types of tumors, although their use is still limited to the diagnosis and prediction of prognosis in meningiomas. To enable targeted molecular therapy informed by gene-panel testing, further studies including matched tumor pair analyses are required to understand the molecular characteristics of meningiomas and develop treatments based on genetic abnormalities.

Abstract Salivary glands develop through epithelial-mesenchymal interactions and are formed through repeated branching. The Crk-associated substrate protein (p130Cas) serves as an adapter that forms a complex with various proteins via integrin and growth factor signaling, with important regulatory roles in several essential cellular processes. We found that p130Cas is expressed in ductal epithelial cells of the submandibular gland (SMG). We generated epithelial tissue-specific p130Cas-deficient (p130Cas Δepi– ) mice and aimed to investigate the physiological role of p130Cas in the postnatal development of salivary glands. Histological analysis showed immature development of granular convoluted tubules (GCT) of the SMG in male p130Cas Δepi– mice. Immunofluorescence staining showed that nuclear-localized androgen receptors (AR) were specifically decreased in GCT cells in p130Cas Δepi– mice. Furthermore, epidermal growth factor-positive secretory granules contained in GCT cells were significantly reduced in p130Cas Δepi– mice with downregulated AR signaling. GCTs lacking p130Cas showed reduced numbers and size of secretory granules, disrupted subcellular localization of the cis-Golgi matrix protein GM130, and sparse endoplasmic reticulum membranes in GCT cells. These results suggest that p130Cas plays a crucial role in androgen-dependent GCT development accompanied with ER-Golgi network formation in SMG by regulating the AR signaling.

Introduction: The role of osteopontin (OPN) following severe injury remains to be elucidated, especially its relationship with type I collagen (encoded by the Col1a1 gene) secretion by newly-differentiated odontoblast-like cells (OBLCs). In this study, we examined the role of OPN in the process of reparative dentin formation with a focus on reinnervation and revascularization after tooth replantation in Opn knockout (KO) and wild-type (WT) mice. Methods: Maxillary first molars of 2- and 3-week-old-Opn KO and WT mice (Opn KO 2W, Opn KO 3W, WT 2W, and WT 3W groups) were replanted, followed by fixation 3–56 days after operation. Following micro-computed tomography analysis, the decalcified samples were processed for immunohistochemistry for Ki67, Nestin, PGP 9.5, and CD31 and in situ hybridization for Col1a1. Results: An intense inflammatory reaction occurred to disrupt pulpal healing in the replanted teeth of the Opn KO 3W group, whereas dental pulp achieved healing in the Opn KO 2W and WT groups. The tertiary dentin in the Opn KO 3W group was significantly decreased in area compared with the Opn KO 2W and WT groups, with a significantly low percentage of Nestin-positive, newly-differentiated OBLCs during postoperative days 7–14. In the Opn KO 3W group, the blood vessels were significantly decreased in area and pulp healing was disturbed with a failure of pulpal revascularization and reinnervation. Conclusions: OPN is necessary for proper reinnervation and revascularization to deposit reparative dentin following severe injury within the dental pulp of erupted teeth with advanced root development.

Abstract Texture has enormous effects on food preferences. The materials used to study texture discrimination also have tastes that experimental animal can detect; therefore, such studies must be designed to exclude taste differences. In this study, to minimize the effects of material tastes, we utilized high- and low-viscosity forms of carboxymethyl cellulose (CMC-H and CMC-L, respectively) at the same concentrations (0.1–3%) for viscosity discrimination tests in rats. In two-bottle preference tests of water and CMC, rats avoided CMC-H solutions above 1% (63 mPa·s) but did not avoid less viscous CMC-L solutions with equivalent taste magnitudes, suggesting that rats spontaneously avoided high viscosity. To evaluate low-viscosity discrimination, we performed conditioned aversion tests to 0.1% CMC, which initially showed a comparable preference ratio to water in the two-bottle preference tests. Conditioning with 0.1% CMC-L (1.5 mPa·s) did not induce aversion to 0.1% CMC-L or CMC-H. However, rats acquired a conditioned aversion to 0.1% CMC-H (3.6 mPa·s) even after latent inhibition to CMC taste by pre-exposure to 0.1% CMC-L. These results suggest that rats can discriminate considerably low viscosity independent of CMC taste. This novel approach for viscosity discrimination can be used to investigate the mechanisms of texture perception in mammals.

Abstract Background Etoposide plus cisplatin (EP) combined with concurrent accelerated hyperfractionated thoracic radiotherapy (AHTRT) is the standard treatment strategy for unresectable limited‐disease (LD) small cell lung cancer (SCLC), which has remained unchanged for over two decades. Based on a previous study that confirmed the non‐inferiority of amrubicin (AMR) plus cisplatin (AP) when compared with EP for extensive‐disease (ED) SCLC, we have previously conducted a phase I study assessing AP with concurrent TRT (2 Gy/time, once daily, 50 Gy in total) for LD‐SCLC therapy. Our findings revealed that AP with concurrent TRT could prolong overall survival to 39.5 months with manageable toxicities. Therefore, we plan to conduct a phase I study to investigate and determine the effect of AP combined with AHTRT, recommended dose (RD), maximum tolerated dose (MTD), and dose‐limiting toxicity (DLT) of AP in patients with LD‐SCLC. Methods Treatment‐naive patients with LD‐SCLC, age between 20 and 75 years, who had a performance status of 0 or 1 and adequate organ functions will be enrolled. For chemotherapy, cisplatin 60 mg/m2/day (day 1) and AMR (day 1 to 3) will be administered with AHTRT (1.5 Gy/time, twice daily, 45 Gy in total). The initial AMR dose is set to 25 mg/m2/day. RD and MTD will be determined by evaluating toxicities. Discussion Based on our previous study, the initial dose of AMR 25 mg/m2 is expected to be tolerated and acceptable. Here, we aim to determine whether treatment with AP and concurrent AHTRT would be an optimal choice with manageable toxicities for LD‐SCLC.

The treatment of World Health Organization (WHO) grades 2 and 3 meningiomas remains difficult and controversial. The pathogenesis of high-grade meningiomas was expected to be elucidated to improve treatment strategies. The molecular biology of meningiomas has been clarified in recent years. High-grade meningiomas have been linked to NF2 mutations and 22q deletion. CDKN2A/B homozygous deletion and TERT promoter mutations are independent prognostic factors for WHO grade 3 meningiomas. In addition to 22q loss, 1p, 14p, and 9q loss have been linked to high-grade meningiomas. Meningiomas enriched in copy number alterations may be biologically invasive. Furthermore, several new comprehensive classifications of meningiomas have been proposed based on these molecular biological features, including DNA methylation status. The new classifications may have implications for treatment strategies for refractory aggressive meningiomas because they provide a more accurate prognosis compared to the conventional WHO classification. Although several systemic therapies, including molecular targeted therapies, may be effective in treating refractory aggressive meningiomas, these drugs are being tested. Systemic drug therapy for meningioma is expected to be developed in the future. Thus, this review aims to discuss the distinct genomic alterations observed in WHO grade 2 and 3 meningiomas, as well as their diagnostic and therapeutic implications and systemic drug therapies for high-grade meningiomas.

Abstract Background Melanoma is a malignant tumor characterized by high proliferation and aggressive metastasis. To address the molecular mechanisms of the proto-oncogene, Rous sarcoma oncogene (Src), which is highly activated and promotes cell proliferation, migration, adhesion, and metastasis in melanoma. Plectin, a cytoskeletal protein, has recently been identified as a Src-binding protein that regulates Src activity in osteoclasts. Plectin is a candidate biomarker of certain tumors because of its high expression and the target of anti-tumor reagents such as ruthenium pyridinecarbothioamide. The molecular mechanisms by which plectin affects melanoma is still unclear. In this study, we examined the role of plectin in melanoma tumor formation. Methods We used CRISPR/Cas9 gene editing to knock-out plectin in B16 mouse melanoma cells. Protein levels of plectin and Src activity were examined by western blotting analysis. In vivo tumor formation was assessed by subcutaneous injection of B16 cells into nude mice and histological analysis performed after 2 weeks by Hematoxylin-Eosin (H&E) staining. Cell proliferation was evaluated by direct cell count, cell counting kit-8 assays, cyclin D1 mRNA expression and Ki-67 immunostaining. Cell aggregation and adhesion were examined by spheroid formation, dispase-based dissociation assay and cell adhesion assays. Results In in vivo tumor formation assays, depletion of plectin resulted in low-density tumors with large intercellular spaces. In vitro experiments revealed that plectin-deficient B16 cells exhibit reduced cell proliferation and reduced cell-to-cell adhesion. Since Src activity is reduced in plectin-deficient melanomas, we examined the relationship between plectin and Src signaling. Src overexpression in plectin knockout B16 cells rescued cell proliferation and improved cell-to-cell adhesion and cell to extracellular matrix adhesion. Conclusion These results suggest that plectin plays critical roles in tumor formation by promoting cell proliferation and cell-to-cell adhesion through Src signaling activity in melanoma cells.

Abstract Regardless of treatment, the clinical progression of neurofibromatosis type 2 (NF2), particularly in terms of hearing, swallowing, and gait, tend to worsen throughout the patients’ lives. We performed a retrospective analysis of functional outcomes in Japanese NF2 patients to predict their functional prognosis. We analyzed genotype–phenotype correlation based on genetic data from a cohort of 57 patients with a mean follow-up of 14.5 ± 6.0 years. Their functional outcomes, including hearing, swallowing, and ambulation, were reviewed. Performing a targeted deep sequencing, germline NF2 mutations were identified in 28 patients (49.1%), and mosaic NF2 was identified in 20 patients (20, 35.0%). The functional preservation period and outcome differed significantly depending on clinical/genetic factors. Among these factors, “Truncating”, “Mosaic”, and “Age of symptom onset ≥ 25” had the most significant effects on functional disability. By applying a combination of an NF2 mutation type/location, and age of symptom onset, we classified different degrees of functional preservation and progression, schwannoma growth rate and total interventions per year per patient. The prediction of detailed functional outcomes in NF2 patients can plan better strategies for life-long disease management and social integration.

Abstract NF2 alteration is the most commonly–found genetic abnormality in meningiomas and is known to initiate events for aggressive-type meningiomas. Whereas the prognosis of meningiomas differs depending on their epigenomic/transcriptomic profile, the effect of NF2 alteration on the prognosis of benign meningiomas is not fully elucidated. This study aimed to probe the importance of NF2 alteration in prognosis of WHO grade I meningiomas. A long-term retrospective follow-up (5.3 ± 4.5 years) study involving 281 consecutive WHO grade I meningioma patients was performed. We assessed tumour recurrence in correlation with extent of resection (EOR), histopathological findings, tumour location, and NF2 alteration. “NF2 meningioma” was defined as meningiomas with presence of NF2 mutation and/or 22q loss. Overall, NF2 meningioma per se was not a predictor of prognosis in the whole cohort; however, it was a predictor of recurrence in supratentorial meningiomas, together with EOR and Ki-67. In a striking contrast, NF2 meningioma showed a better prognosis than non-NF2 meningioma in infratentorial lesion. Supratentorial NF2 meningiomas had higher Ki-67 and forkhead box protein M1 expression than those of others, possibly explaining the worse prognosis in this subtype. The combination of NF2 alteration, high Ki-67 and supratentorial location defines subgroup with the worst prognosis among WHO grade I meningiomas. Clinical connotation of NF2 alteration in terms of prognosis of WHO grade I meningioma differs in an opposite way between supratentorial and infratentorial tumors. Integrated anatomical, histopathological, and genomic classifications will provide the best follow-up schedule and proactive measures.