Pathological and biological significance of the specific glycan, TRA-1-60, on aggressive gastric adenocarcinoma.

The glycans form a unique complex on the surface of cancer cells and play a pivotal role in tumor progression, impacting proliferation, invasion, and metastasis. TRA-1-60 is a glycan that was identified as a critical marker for the establishment of fully reprogrammed inducible pluripotent stem (iPS) cells. Its expression has been detected in multiple cancer tissues, including embryonal carcinoma, prostate cancer, and pancreatic cancer, but the biological and pathological characterization of TRA-1-60-expressing tumor cells still remains unclear within various types of malignancies. Here, we report the biological characteristics of TRA-1-60-expressing gastric cancer cells, especially those with its cell surface expression, and the therapeutic significance of targeting TRA-1-60. The cells with cell membrane expression of TRA-1-60 were mainly observed in the invasive area of patient gastric cancer tissues and correlated with advanced stages of the disease based on histopathological and clinicopathological analyses. In vitro analysis using a scirrhous gastric adenocarcinoma line, HSC-58, which highly expresses TRA-1-60 on its plasma membrane, revealed increased stress-resistant mechanisms, supported by the upregulation of glutathione synthetase (GSS) and NCF-1 (p47phox) via lipid-ROS regulatory pathways, as detected by RNA-seq analysis followed by oxidative stress gene-profiling. Our in vivo therapeutic study using the TRA-1-60-targeting antibody-drug conjugate (ADC), namely Bstrongomab conjugated Monomethyl auristatin E (MMAE), showed robust efficacy in a mouse model of peritoneal carcinomatosis induced by intraperitoneal xenograft of HSC-58, by markedly reducing massive tumor ascites. Thus, targeting the specific cell surface glycan, TRA-1-60, shows significant therapeutic impact in advanced-stage gastric cancers. (243 words).

CD14+ Dendritic-Shaped Cells Functioning as Dendritic Cells in Rheumatoid Arthritis Synovial Tissues.

OBJECTIVE: We previously reported that CD14+ dendritic-shaped cells exhibit a dendritic morphology, engage in pseudo-emperipolesis with lymphocytes, and express CD90 in the perivascular areas of rheumatoid arthritis (RA) synovial tissues. However, it remains unclear whether these CD14highCD90intermediate(int) cells function as dendritic cells. In this study, we investigated the dendritic cell-differentiation potential of CD14highCD90int cells. METHODS: The localization and number of CD14highCD90int cells in RA synovial tissues and peripheral blood were examined. The dendritic cell-differentiation potential of CD14highCD90int cells was examined by measuring interleukin-6 and tumor necrosis factor-α levels in the supernatant and CD83 and human leukocyte antigen (HLA)-DR expression in the cells after induction of dendritic cell differentiation. Synovial cells were co-cultured with lymphocytes, and the activation of these cells was examined. RESULTS: CD14highCD90int cells were abundant in RA synovial tissues, including the sublining layer and the pannus areas. Patients with untreated and active RA had significantly higher percentages of CD14highCD90int cells in the peripheral blood and synovial tissues. In RA synovial cells, inflammatory cytokine levels increased with dendritic cell-differentiation culture, but CD83 and HLA-DR expression were significantly increased in the CD14highCD90int cell group. When co-cultured with lymphocytes, cell numbers and inflammatory cytokine levels significantly increased in both groups of synovial cells after dendritic cell induction. CONCLUSION: CD14+ cells migrate and spread from the circulating blood to RA synovial tissues while expressing CD90, and CD14highCD90int cells in contact with lymphocytes differentiate into HLA-DR+ dendritic cells, which contribute to chronic inflammation in RA.

Possible involvement of DExD/H box helicase 60 in synovial inflammation of rheumatoid arthritis: role of toll-like receptor 3 signaling.

BACKGROUND: Innate immunity is known to be implicated in the etiology of synovitis in rheumatoid arthritis (RA). However, details of the molecular mechanisms have not been fully clarified. DExD/H-box helicase 60 (DDX60), a putative RNA helicase, is of consequence in anti-viral innate immune reactions followed by inflammation. Although DDX60 is involved in the pathogenesis of autoimmune diseases such as systemic lupus nephritis, the role of DDX60 in RA has not been elucidated. The objective of this study was to examine the expression and the role of DDX60 in RA synovial inflammation. METHODS AND RESULTS: DDX60 protein expression was investigated by immunohistochemistry in synovial tissues resected from 4 RA and 4 osteoarthritis (OA) patients. We found that synovial DDX60 expression was more intense in RA than in OA. Treatment of human rheumatoid fibroblast-like synoviocytes in culture with polyinosinic-polycytidylic acid, a Toll-like receptor 3 (TLR3) ligand, increased DDX60 protein and mRNA expression. A knockdown experiment of DDX60 using RNA interference revealed a decrease in the expression of poly IC-induced C-X-C motif chemokine ligand 10 (CXCL10) which induces lymphocyte chemotaxis. CONCLUSIONS: The synovial DDX60 was more expressed in RA patients than in OA. In human RFLS, DDX60 stimulated by TLR3 signaling affected CXCL10 expression. DDX60 may contribute to synovial inflammation in RA.

Inflammatory Rhabdomyoblastic Tumor: Clinicopathologic and Molecular Analysis of 13 Cases.

Inflammatory rhabdomyoblastic tumors (IRMTs) are newly recognized skeletal muscle tumors with uncertain malignant potential. We investigated 13 IRMTs using clinicopathologic, genetic, and epigenetic methods. The cohort included 7 men and 6 women, aged 23 to 80 years (median, 50 years), of whom 2 had neurofibromatosis type 1. Most tumors occurred in the deep soft tissues of the lower limbs, head/neck, trunk wall, and retroperitoneum/pelvis. Two tumors involved the hypopharyngeal submucosa as polypoid masses. Eight tumors showed conventional histology of predominantly spindled cells with nuclear atypia, low mitotic activity, and massive inflammatory infiltrates. Three tumors showed atypical histology, including uniform epithelioid or plump cells and mitotically active histiocytes. The remaining 2 tumors demonstrated malignant progression to rhabdomyosarcoma; one had additional IRMT histology and the other was a pure sarcoma. All 11 IRMTs without malignant progression exhibited indolent behavior at a median follow-up of 43 months. One of the 2 patients with IRMTs with malignant progression died of lung metastases. All IRMTs were positive for desmin and PAX7, whereas myogenin and MyoD1 were expressed in a subset of cases. Targeted next-generation sequencing identified pathogenic mutations in NF1 (5/8) and TP53 (4/8). All TP53 mutations co-occurred with NF1 mutations. TP53 variant allele frequency was much lower than that of NF1 in 2 cases. These tumors showed geographic (subclonal) strong p53 immunoreactivity, suggesting the secondary emergence of a TP53-mutant clone. DNA methylation-based copy number analysis conducted in 11 tumors revealed characteristic flat patterns with relative gains, including chromosomes 5, 18, 20, 21, and/or 22 in most cases. Widespread loss of heterozygosity with retained biparental copies of these chromosomes was confirmed in 4 tumors analyzed via allele-specific profiling. Based on unsupervised DNA methylation analysis, none of the 11 tumors tested clustered with existing reference entities but formed a coherent group, although its specificity warrants further study.

A Case of Disseminated Non-tuberculous Mycobacteriosis with a Skull Lesion Controlled by Resection after Exacerbation during Antimicrobial Chemotherapy in a Patient Positive for Anti-IFN-γ-neutralizing Autoantibodies.

A 48-year-old man presented with a fever and back pain and was referred to our hospital with multiple bone destruction and abscess formation. A sputum examination revealed Mycobacterium intracellulare, and pathological findings revealed an indistinct granuloma and acid-fast bacilli, leading to a diagnosis of disseminated nontuberculous mycobacteriosis. Anti-interferon-γ-neutralizing autoantibodies were detected in the serum, and acquired immunodeficiency was suspected to be the etiology. Antimicrobial chemotherapy was initiated, and the lesions generally regressed. However, only the skull lesions worsened, requiring local resection to control the disease. Currently, the patient is continuing to receive drug therapy with good disease control after debridement.

Small Cell Neuroendocrine Carcinoma of the Parotid Gland: An Uncommon Example of Incompletely Encapsulated Tumor Including S100 Protein-Positive Clear Cells.

Small cell carcinoma is rare in salivary glands and has recently been termed small cell neuroendocrine carcinoma. We herein describe an uncommon example arising in the parotid gland. The patient was a 75 yearold Japanese male who had swelling in the right parotid area. He underwent a superficial lobectomy and, after a histological diagnosis was made, a total parotidectomy. Histologically, the tumor had a thick hyalinized capsule that was incomplete, beyond which the tumor invaded into the surrounding parotid parenchyma. The tumor consisted of typical small basophilic cells intermingled with bland clear cells, between which a gradual transition was observed both inside and outside the capsule. Small basophilic cells were immunoreactive for chromograninA as well as synaptophysin, while clear cells were positive for S100 protein. The Ki-67 labeling rate reached 30-40% at the high points of small basophilic cells, but clear cells were minimally labelled. The present case was considered a dedifferentiated carcinoma of the parotid gland, possibly with acinic cell carcinoma as a precursor. This tumor could also be considered a "mixed exocrine-endocrine carcinoma," which may explain the histogenesis of neuroendocrine carcinomas in non-endocrine organs that are not included in the diffuse (dispersed) neuroendocrine system, such as the parotid gland.

Spontaneous remission of skull Langerhans cell histiocytosis that had developed by repeated head injury: illustrative case.

BACKGROUND: Langerhans cell histiocytosis (LCH) was previously characterized as the proliferation of Langerhans-type histiocytes with a wide range of clinical presentations that arise mostly in children. The typical presentation is a gradually enlarging, painless skull mass. Rapid clinical deterioration is rare. OBSERVATIONS: A 3-year-old boy who had incurred a right frontal impact head injury demonstrated no apparent neurological deficits. He subsequently bruised the same region multiple times. The right frontal swelling gradually increased over the course of 6 days after the initial injury. Skull radiography showed no bony lesion. The same site enlarged markedly 12 days after the initial injury. Magnetic resonance imaging revealed a frontal bony tumorous lesion associated with multiple subcutaneous cystic mass lesions. The patient underwent open biopsy of the skull lesion and evacuation of the subcutaneous lesions. Histopathological examination confirmed the diagnosis of LCH. Immunohistochemical evaluation revealed positivity for CD1a and langerin and no immunopositivity for BRAF V600E. The skull lesion spontaneously disappeared 30 days after the biopsy without recurrence. LESSONS: Physicians should be aware of this rare clinical manifestation of LCH that developed by a repeat head injury.

Vasopressin Expressed in Hypothalamic CRF Neurons Causes Impaired Water Diuresis in Secondary Adrenal Insufficiency.

Patients with secondary adrenal insufficiency can present with impaired free water excretion and hyponatremia, which is due to the enhanced secretion of vasopressin (AVP) despite increased total body water. AVP is produced in magnocellular neurons in the paraventricular nucleus of the hypothalamus (PVH) and supraoptic nucleus and in parvocellular corticotropin-releasing factor (CRF) neurons in the PVH. This study aimed to elucidate whether magnocellular AVP neurons or parvocellular CRF neurons coexpressing AVP are responsible for the pathogenesis of hyponatremia in secondary adrenal insufficiency. The number of CRF neurons expressing copeptin, an AVP gene product, was significantly higher in adrenalectomized AVP-floxed mice (AVPfl/fl) than in sham-operated controls. Adrenalectomized AVPfl/fl mice supplemented with aldosterone showed impaired water diuresis under ad libitum access to water or after acute water loading. They became hyponatremic after acute water loading, and it was revealed under such conditions that aquaporin-2 (AQP2) protein levels were increased in the kidney. Furthermore, translocation of AQP2 to the apical membrane was markedly enhanced in renal collecting duct epithelial cells. Remarkably, all these abnormalities observed in the mouse model for secondary adrenal insufficiency were ameliorated in CRF-AVP-/- mice that lacked AVP in CRF neurons. Our study demonstrates that CRF neurons in the PVH are responsible for the pathogenesis of impaired water excretion in secondary adrenal insufficiency.

Involvement of the genus Corynebacterium in the pathogenesis of pigmented intratarsal keratinous cyst.

Intratarsal keratinous cyst (IKC) is a benign cystic lesion of the eyelid that retains keratin flakes. IKCs are usually yellow to white cystic lesions but rarely become brown or gray-blue, making clinical diagnosis difficult. The mechanisms by which dark brown pigments are generated in pigmented IKC are unclear. The authors report a case of pigmented IKC that had melanin pigments within the lining of the cyst wall and within the cyst. Focal infiltrates of lymphocytes were observed in the dermis, particularly beneath the cyst wall in areas with more melanocytes and intense melanin deposition. These pigmented parts faced bacterial colonies inside the cyst, which were identified to be Corynebacterium species in a bacterial flora analysis. The pathogenesis of pigmented IKC in relation to inflammation and bacterial flora is discussed.

Rescue computed tomography-guided drainage of a giant mediastinal mature teratoma causing airway obstruction prior to surgical resection: a case report.

BACKGROUND: Giant mediastinal mature teratomas may cause airway obstruction or decreased venous return due to the mass effect. Preoperative stabilization of the respiratory and circulatory systems is important for perioperative management to safely perform surgery, including general anesthesia. However, to the best of our knowledge, there are only a few reports regarding the preoperative computed tomography (CT)-guided drainage of mediastinal tumors. CASE PRESENTATION: A 30-year-old woman was admitted to the emergency room with sudden dyspnea. CT findings revealed a giant cystic mass in the anterior mediastinum compressing the trachea and the right main bronchus. The patient was intubated and CT-guided drainage of the fluid content of the cyst was performed to decompress the airway obstruction. Thereafter, the mediastinal tumor was resected during elective surgery and pathologically diagnosed as a mature teratoma. CONCLUSIONS: Rescue preoperative CT-guided drainage of a giant mediastinal mature teratoma allowed safe general anesthesia and surgery by releasing the airway obstruction.

Spindle Cell Variant of Ameloblastic Carcinoma: Another Example in a Japanese Male.

Spindle cell variant of ameloblastic carcinoma (SpCAC) is a rare subtype of ameloblastic carcinoma. Herein, we describe an additional case of SpCAC of the mandible of a 76-year-old Japanese male. We discuss diagnostic problems we encountered in this case, focusing on unusual expression of myogenic/myoepithelial markers, such as smooth muscle actin and calponin.

Granulomatosis with Polyangiitis following COVID-19 in an Adolescent.

Granulomatosis with polyangiitis (GPA) is a systemic disease that causes vasculitis in various organs. Although the mechanism of pathogenesis remains unclear, infection has been reported to be a causative factor. We herein report a case of GPA that developed following coronavirus disease 2019 (COVID-19) in an adolescent girl. One month after contracting mild COVID-19, the patient had facial allodynia, a fever, and weight loss and was admitted for multiple nodular shadows on a chest roentgenogram. GPA was diagnosed based on pathological findings of the lung and nasal mucosal biopsies. She received methylprednisolone and rituximab, and her symptoms and radiological findings improved.

Utility of arterial spin labeling for objective assessment of intratumoral microvessels in diffuse hemispheric glioma, H3 G34R-mutant: A case report and literature review.

Imaging findings of diffuse hemispheric glioma H3 G34-mutant (DHG, H3 G34m), a new variant of glioma under the World Health Organization classification, have recently been vigorously debated. Here, we report a case of DHG, H3 G34m in which objective assessments of intratumoral microvessels using arterial spin labeling (ASL) were useful for preoperative diagnosis, selection of anti-tumor drugs, and tracking therapeutic responses. The patient was a 34-year-old woman who presented with weakness in the left arm. Preoperative magnetic resonance imaging (MRI) showed no specific findings of hyperintensity on fluid-attenuated inversion recovery imaging and faint enhancement on T1-weighted imaging with contrast media in the tumor. However, ASL showed a convincing finding of high blood flow in the entire tumor, allowing identification of the tumor as malignant glioma. Tumor specimens obtained from biopsy showed that the tumor comprised low-differentiated tumor cells, abundant histiocytes, and highly dense microvessels. Immunohistochemical findings such as positive findings for H3 G34R and p53, and negative findings for IDH-1, ATRX, and OLIG2 led to the diagnosis of DHG, H3 G34m. Based on findings of hyperperfusion on ASL and detection of vascular endothelial growth factor (VEGF), we administered the anti-VEGF antibody bevacizumab. The tumor shrank significantly but remained. However, the residual tumor showed hypoperfusion on ASL, strongly suggesting tumor remission. Objective assessments of blood flow using ASL are useful in clinical practice for patients with DHG, H3 G34 showing non-specific findings on conventional MRI.

Establishment of isogenic induced pluripotent stem cells with or without pathogenic mutation for understanding the pathogenesis of myeloproliferative neoplasms.

Identification and functional characterization of disease-associated genetic traits are crucial for understanding the pathogenesis of hematologic malignancies. Various in vitro and in vivo models, including cell lines, primary cells, and animal models, have been established to examine these genetic alterations. However, their nonphysiologic conditions, diverse genetic backgrounds, and species-specific differences often limit data interpretation. To evaluate somatic mutations in myeloproliferative neoplasms (MPNs), we used CRISPR/Cas9 combined with the piggyBac transposon system to establish isogenic induced pluripotent stem (iPS) cell lines with or without JAK2V617F mutation, a driver mutation of MPNs. We induced hematopoietic stem/progenitor cells (HSPCs) from these iPS cells and observed phenotypic differences during hematopoiesis using fluorescence-activated cell sorting analysis. HSPCs with pathogenic mutations exhibited cell-autonomous erythropoiesis and megakaryopoiesis, which are hallmarks in the bone marrow of patients with MPNs. Furthermore, we used these HSPCs as a model to validate therapeutic compounds and showed that interferon alpha selectively inhibited erythropoiesis and megakaryopoiesis in mutant HSPCs. These results demonstrate that genome editing is feasible for establishing isogenic iPS cells, studying genetic elements to understand the pathogenesis of MPNs, and evaluating therapeutic compounds against MPNs.

Dysembryoplastic Neuroepithelial Tumor of the Infratentorial Multiple Lesions: A Case Report and Review of the Literature.

A dysembryoplastic neuroepithelial tumor (DNT) is a benign neoplasm that usually occurs in the supratentorial cerebral cortex. Here, we report a rare case of an infratentorial DNT in a 42-year-old woman who presented with dizziness and a gait disturbance. Magnetic resonance imaging of the lesion demonstrated hyperintensity on T2-weighted images and hypointensity on T1-weighted images of the left cerebellar hemisphere with a multifocal lesion. Macroscopically, the lesion appeared soft, avascular, and slightly torose at the cortical surface. Histologically, dysplastic disorganization of the cortex and floating neurons were observed. The pathological and immunochemical features of this case agree with the diagnosis of a DNT. The lesion partially included cortical heterotopia, which is a novel observation in an infratentorial DNT. On the basis of the previous reports, we discussed the surgical resection of the infratentorial DNT.

Telomere length was associated with grade and pathological features of meningioma.

Telomeres are tandem repeats of the TTAGGG sequence at chromosomal ends and afford protection against chromosomal instability. To investigate the contribution of telomere dysfunction in meningiomas, here we estimate the associations between telomere length, tumor grade, and proliferation index in a series of 14 archived samples, using quantitative-fluorescence in situ hybridization, Ki67 immunostaining, and pathological analysis. The number of mitoses per 10 high-power fields (HPF) and Ki67 index was higher in grade III cases than in grade I or grade II cases. Telomere length was negatively associated with both the number of mitoses/10HPF and Ki67 index. Meningioma cases with atypical mitosis, a morphological marker of chromosomal instability, exhibited shortened telomeres. Among telomere-shortened meningioma cases, 40% were grade I, 20% were grade II, and 100% were grade III. In grade I or II meningiomas, shortened telomeres lacked high proliferation activity and atypical mitosis. In conclusion, telomere shortening might be pivotal in the development of high-grade meningioma. Analysis of telomere length might be a selective marker for meningiomas with high-grade malignant potential.

Association of CD90 Expression by CD14+ Dendritic-Shaped Cells in Rheumatoid Arthritis Synovial Tissue With Chronic Inflammation.

OBJECTIVE: CD14+ dendritic-shaped cells show a dendritic morphology under the electron microscopy and engage in a pseudoemperipolesis phenomenon with lymphocytes. CD90 has been used as a marker of a major subset of fibroblast-like synoviocytes in rheumatoid arthritis (RA). In this study, we investigated the significance of CD90 expression in CD14+ dendritic-shaped cells and its correlation with RA chronic inflammation. METHODS: Double immunofluorescence staining for CD14 and CD90 was performed in the collected tissues, including 12 active RA synovial tissues. The localization of CD14+ CD90+ cells, the percentages of CD14+ CD90+ cells and vascular areas, the degree of synovitis, and clinical data were investigated. Furthermore, CD14+ CD90+ cells analyzed by flow cytometry (CD14high CD90intermediate (int) cells) were sorted from RA synovial cells, and we examined their potential to differentiate into dendritic cells. RESULTS: Double immunofluorescence staining showed that CD14+ CD90+ cells were abundant in RA synovial tissues. The percentages of CD14+ CD90+ cells and vascular areas correlated with some of the Krenn synovitis scores, but neither showed a strong correlation with RA disease activity parameters. Flow cytometry analysis indicated that CD14high CD90int cells were more abundant in both peripheral blood samples and synovial tissues in patients with active RA. CD14high CD90int cells were more likely to differentiate into dendritic cells in vitro. CONCLUSION: CD14+ dendritic-shaped cells expressed CD90 in the perivascular areas of RA synovial tissues. These findings suggest that CD14+ CD90+ dendritic-shaped cells migrate from the peripheral blood to the synovial tissue, the site of inflammation, and may contribute to the chronic inflammation of RA as dendritic progenitor cells.

Clinicopathological risk factors for a poor prognosis of primary central nervous system lymphoma in elderly patients in the Tohoku and Niigata area: a multicenter, retrospective, cohort study of the Tohoku Brain Tumor Study Group.

Clinicopathological risk factors for a poor prognosis were investigated in elderly patients with malignant lymphoma of the central nervous system. A total of 82 pathologically confirmed, CD20-positive, diffuse large B-cell lymphoma patients aged 71 years or older who underwent therapeutic intervention in the Tohoku and Niigata area in Japan were retrospectively reviewed. A univariate analysis was performed by the log-rank test using the Kaplan-Meier method. A Cox proportional hazards model was used for multivariate analysis of risk factors. Of the 82 patients, 39 were male and 43 were female, and their median age at onset was 75 years. At the end of the study, there were 34 relapse-free patients (41.5%), 48 relapse cases (58.5%), median progression-free survival was 18 months, and median overall survival (OS) was 26 months; there were 41 deaths and 41 survivors. Multivariate analysis of median OS showed that Karnofsky Performance Status less than 60% 3 months after treatment (p = 0.022, hazard ratio (HR) = 2.591) was the clinical risk factor, and double expressor lymphoma (p = 0.004, HR = 3.163), expression of programmed death-ligand 1 in tumor infiltrating lymphocytes or tumor-associated macrophages (p < 0.001, HR = 5.455), and Epstein-Barr virus infection (p = 0.031, HR = 5.304) were the pathological risk factors.

The Real-World status and risk factors for a poor prognosis in elderly patients with primary central nervous system malignant lymphomas: a multicenter, retrospective cohort study of the Tohoku Brain Tumor Study Group.

BACKGROUND: Elderly patients with primary central nervous system malignant lymphoma (EL-PCNSL) may not be given sufficient treatment due to their poor pre-treatment Karnofsky Performance Status (KPS) and comorbidities. Therefore, a retrospective, cohort study was performed to evaluate risk factors associated with a poor prognosis of EL-PCNSL in the Tohoku Brain Tumor Study Group. METHODS: Patients aged ≥ 71 years with PCNSL were enrolled from eight centers. Univariate analysis was performed with the log-rank test. A Cox proportional hazards model was used for multivariate analysis. RESULTS: Three of the total 142 cases received best supportive care (BSC). Treatment was given to 30 cases without a pathological diagnosis, 3 cases with cerebrospinal fluid (CSF) cytology, and 100 cases with a pathological diagnosis. After confirmation of no differences in progression-free survival (PFS) and overall survival (OS) between the group treated without pathology and the groups diagnosed by pathology or CSF cytology and between median age ≥ 76 years and < 76 years, a total of 133 patients were studied. The median pre-treatment KPS was 50%. Median PFS and median OS were 16 and 24 months, respectively. Risk factors associated with poor prognosis on Cox proportional hazards model analysis were pre-treatment cardiovascular disease and central nervous system disease comorbidities, post-treatment pneumonia and other infections, and the absence of radiotherapy or chemotherapy. CONCLUSIONS: Pre-treatment comorbidities and post-treatment complications would affect the prognosis. Radiation and chemotherapy were found to be effective, but no conclusions could be drawn regarding the appropriate content of chemotherapy and whether additional radiotherapy should be used.