Jing Zhang

Background: Ewing’s sarcoma (EWS) is a highly aggressive malignant tumor that originates from bone or soft tissue. To date, there is no established prognostic model for EWS tumor. This study aims to identify prognostic genes and develop a predictive model associated with fatty acid metabolism in EWS using bioinformatics analysis. Results: We analyzed the GSE17679 dataset and identified 25 differentially expressed genes related to fatty acid metabolism in EWS. A risk model composed of ACADM, ADH5, ACSL1, ELOVL4, ECI1, PPT1, and ACOT7 gene signatures was constructed. The AUC values at 3 and 5 years were both ≥0.7, indicating good predictive accuracy. GSVA analysis revealed significant differences in fatty acid metabolism pathway enrichment between high- and low-risk groups. Differential genes were primarily enriched in pathways such as fatty acid oxidation, lipid oxidation, lipid modification, and fatty acid degradation. Immune infiltration analysis showed significant differences in memory B cells, activated NK cells, and neutrophils between the two groups. Additionally, significant differences were observed in the expression of immune checkpoints such as HAVCR2 and LDHB. Immunohistochemistry and survival analysis further demonstrated that the expression of PPT1 and ACOT7 proteins was associated with the progression-free survival of EWS patients. Conclusions: We successfully constructed a prognostic model for EWS related to fatty acid metabolism genes. PPT1 and ACOT7 may serve as promising predictors for EWS prognosis.

Background: As the only humanized monoclonal antibody against receptor activator of nuclear factor-κB ligand (RANKL) for giant cell tumour of bone (GCTB) therapy, denosumab has limited antitumour effect on neoplastic stromal cells. Nevertheless, its mechanism of action has not yet been clarified. A previous study has revealed that p62 may play an important role in the antitumour activity of denosumab. Objective: The study aimed to investigate if the mechanism by which denosumab inhibits GCTB neoplastic stromal cells growth is via p62 modulation and other related mechanisms. Methods: p62 expression before and after denosumab therapy was analysed by RT‒qPCR, western blot, ELISA, and immunohistochemical assays. Two primary neoplastic stromal cells were isolated from fresh GCTB tumour tissue (L cell) and metastatic tissue (M cell). Cell proliferation, migration, apoptosis, and autophagy were investigated in p62 knockdown neoplastic stromal cells transfected by short hairpin RNA lentivirus in vitro. Tumor growth was evaluated in the chick chorioallantoic membrane model in vivo. Results: p62 expression was found to be downregulated following denosumab therapy. The patients with a decrease in p62 expression had lower recurrence-free survival rates. The proliferation of M cells was not inhibited by denosumab therapy, but it was restored by p62 knockdown. Moreover, p62 knockdown inhibited tumour growth in vivo. Denosumab induced M cell apoptosis and arrested the cell cycle at the G1/G0 transition and these effects were also enhanced by p62 knockdown. Autophagic flux assays revealed p62 modulation to be dependent on autophagy following denosumab incubation. Conclusion: Denosumab induced neoplastic stromal cells apoptosis via p62 downregulation dependent on autophagy pathway. The combination of p62 and RANKL knockdown might be a better strategy than RANKL knockdown alone for GCTB targeted therapy.

Masquelet technique demonstrated superiority in reconstructing long bone defect after trauma or infection. However, reports in foot tumor were rare. A 24-year-old male diagnosed with calcaneal chondroblastoma who had a defect of calcaneal after intralesional curettage. We reconstructed the defect by Masquelet technique. This is the first case as far as we know that reported Masquelet technique for calcaneal tumor. The technique to treat irregular bone defects after operation can be considered in other similar situations.

Background: Postoperative bone defect is difficult to reconstruct after resection or curettage of bone tumor, especially for some tumor located in ankle and foot. Induced membrane of Masquelet technique demonstrated superiority in reconstructing bone defect after injury, but its safety and validity in foot tumor were not clear. Case presentation: A 24-year-old male patient complained pain in the right heel for more than twoyears. CT scan showed a low density lesion with cortical erosion of calcaneus body. The pathological results indicatedchondroblastoma by biopsy. The first step, tumor was totally removed by curettage. High speed grinding and 95% alcohol inactivation killed the residual tumor cells. Cortex defect of calcaneal was about 8cm in longest diameter. Bone cement (PMMA) spacer was packed into the defect and fixed with two crossedKirschner wires for six weeks. The second step, bone cement was removed. Crossed autologous fibular segments shoring with two screws and allogeneic cancellous graft were filled into the cavity to reconstruct the bone defect. For near one year follow up, the calcaneal bone fusion with graft was good, and foot function recovered satisfactorily. Conclusions: we offered a new reconstructive choice for calcaneal defect after tumor curettage. The safety and validity of Masquelet technique is worth trying in future by more clinical cases.

Induced pluripotent stem cells (iPSCs) are a new potential cure for diabetes, characterised by a capacity for self-renewal and differentiation to pancreatic islet beta cells, which secrete insulin and rebuild blood glucose balance. The safety and validity of iPSC-derived cell therapy for diabetes remain controversial. Teratoma formation arising from undifferentiated stem cells is a serious risk, but clinical reports of this phenomenon are rare. Here, we report a distinctive case of immature teratoma after the patient underwent iPSC-derived cell therapy for diabetes in another hospital, and he wastreated in our soft tissue sarcoma centre. The patient received islet beta cell injection, in which the cells were differentiated from autologous iPSCs, into the deltoid muscle. Two months later, a mass located in the injection area was detected and presented with enlarged axillary lymph nodes. Here, we present the clinical, radiological, and pathological features of this immature teratoma. Distinct from typical immature teratomas, this tumour was characterised by rapid growth and local lymph node metastasis. The tumour did not respond to typical chemotherapy regimens for immature teratomas. Magnetic resonance imaging (MRI) showed heterogeneous enhancement and a rich blood supply to the tumour. Histopathology revealed immature endoderm, mesoderm, and ectoderm tissues composed of osseous, cartilaginous, vascular, and adenoid tissues, which have more cellular atypia than typical teratomas. Staining for both OCT4 and SOX2 was positive in the tumour cell nucleus as revealed by immunofluorescence assay; however, insulin staining was negative. Next-generation sequencing showed many missense mutations, but abnormal gene rearrangement, defects, or changes in copy numbers were not observed. In conclusion, more attention should be given to teratoma formation after iPSC-derived cell therapy for diabetes, because these tumours are more aggressive than typical teratomas. The safety and validity of iPSC-derived cell therapy for diabetes should be explored further in standardised clinical trials.

Background: As a new potential cure for diabetes, induced pluripotent stem cells (iPSCs) are characterized by self-renewal capacity and the ability to differentiate into pancreatic islet β-cells, which will hopefully secrete insulin and rebuild blood glucose balance. The safety and validity of iPSC treatment for diabetes remain controversial. One of the most serious risks is teratoma formation arising from undifferentiated stem cells, but clinical reports are rare.Case presentation: Here, we report a distinctive case of immature teratoma after iPSC treatment for diabetes, which was accidentally treated in our soft tissue sarcoma centre. The patient received islet β-cell injection, in which the cells were differentiated from autologous iPSCs, into the deltoideus muscle. Two months later, a mass located in the injection area formed with enlarged axillary lymph nodes. Here, we present the different clinical, radiological and pathological features of this immature teratoma in detail. Distinctive from typical immature teratomas, the tumour was characterized by rapid growth and local lymph node metastasis. The tumour was not sensitive to typical chemotherapy regimens for immature teratomas. MRI scanning showed heterogeneous enhancement and a rich blood supply to the tumour. The histopathology showed immature endoderm, mesoderm and ectoderm tissues composed of osseous, cartilaginous, vascular and adenoid tissues, which have more cellular atypia than typical teratomas. Staining for both Oct-4 and Sox-2 was positive in the tumour cell nucleus by immunofluorescence assay, but insulin staining was negative. Next-generation sequencing showed many missense mutations, but abnormal gene rearrangement or copy number was not observed.Conclusions: More attention should be given to teratoma formation after iPSC treatment for diabetes, which is more aggressive than typical teratomas. The safety and validity of iPSC treatment for diabetes should be confirmed by more standardized clinical trials.

As a new potential cure for diabetes, induced pluripotent stem cells (iPSCs) are characterized by self-renewal capacity and the ability to differentiate into pancreatic islet β-cells, which will hopefully secrete insulin and rebuild blood glucose balance. The safety and validity of iPSC treatment for diabetes remain controversial. One of the most serious risks is teratoma formation arising from undifferentiated stem cells, but clinical reports are rare. Here, we report a distinctive case of immature teratoma after iPSC treatment for diabetes, which was accidentally treated in our soft tissue sarcoma centre. The patient received islet β-cell injection, in which the cells were differentiated from autologous iPSCs, into the deltoideus muscle. Two months later, a mass located in the injection area formed with enlarged axillary lymph nodes. Here, we present the different clinical, radiological and pathological features of this immature teratoma in detail. Distinctive from typical immature teratomas, the tumour was characterized by rapid growth and local lymph node metastasis. The tumour was not sensitive to typical chemotherapy regimens for immature teratomas. MRI scanning showed heterogeneous enhancement and a rich blood supply to the tumour. The histopathology showed immature endoderm, mesoderm and ectoderm tissues composed of osseous, cartilaginous, vascular and adenoid tissues, which have more cellular atypia than typical teratomas. Staining for both Oct-4 and Sox-2 was positive in the tumour cell nucleus by immunofluorescence assay, but insulin staining was negative. Next-generation sequencing showed many missense mutations, but abnormal gene rearrangement or copy number was not observed. In conclusion, more attention should be given to teratoma formation after iPSC treatment for diabetes, which is more aggressive than typical teratomas. The safety and validity of iPSC treatment for diabetes should be confirmed by more standardized clinical trials.

P62 protein has been implicated in bone metastasis and is a multifunctional adaptor protein usually correlated with autophagy. Herein, we investigated p62 expression and its prognostic significance in bone metastasis of lung adenocarcinoma, and analyzed whether the mechanism involved depends on autophagy. mRNA and protein expression of p62, LC3B and Beclin 1 were detected by reverse transcription-quantitative PCR and western blotting, respectively, in fresh bone metastasis tissues (n=6 cases) and normal cancellous bone tissues (n=3 cases). The association between p62 and LC3B expression and patient prognosis was subsequently analyzed in 62 paraffin-embedded bone metastasis specimens by immunohistochemistry assay. Small interfering RNA (siRNA) was employed to downregulate p62 expression in SPC-A-1 and A549 cells. Cell proliferation and migration ability were tested by CCK8, CCF and Transwell assays respectively. Autophagy was induced by Rapamycin or inhibited by Atg 7 knockout/Chloroquine in A549 cells and p62 and LC3II/I expression were analyzed. After subcutaneous inoculation or intracardial injection of A549 cells into nude mice, the effect of p62 downregulation in vivo was analyzed by histopathological examination. The results showed that p62, LC3B and Beclin 1 mRNA and protein were all overexpressed in bone metastasis tissues (all P3), shorter overall survival rates and shorter progression free survival rates compared with patients having lower p62 expression (P=0.014, P=0.003, P=0.048, respectively). Cox regression analysis identified p62 expression as an independent prognostic indicator of overall survival of patients with bone metastasis (P=0.007). In vitro p62 downregulation inhibited SPC-A-1 and A549 cells migration but had no effect on cell proliferation. After autophagy induction or inhibition, p62 expression involved in autophagy flux and changed inconsistently according to the switch of LC3I to LC3II in different autophagy conditions. In vivo p62 downregulation had no effect on growth of subcutaneous tumor. Lung or bone metastasis lesion was not found in all mice model. These findings suggested that p62 overexpression promotes tumor cell invasion out of LC3-dependent autophagy, which could be used a potential prognostic biomarker and therapeutic target for bone metastasis of lung adenocarcinoma.

Giant cell tumor of bone (GCTB) is an intermediate (locally aggressive) bone tumor with a recurrence rate of >30% following surgery. GCTB recurrence is ultimately due to the proliferation of neoplastic stromal (NS) cells. However, the precise mechanism underlying the regulation of NS cell proliferation remains unknown. p62 protein is a multifunctional adaptor protein that exerts a positive role in bone tumors and metabolic bone diseases. In the present study, the mRNA and protein expression levels of p62 were detected by reverse transcription‑quantitative PCR and western blotting, respectively, in 8 paired fresh GCTB tumor tissues and adjacent normal cancellous bone tissues. The association between p62 expression level and patient prognosis was subsequently analyzed in 54 paraffin‑embedded tumor specimens by immunohistochemistry assay. NS cells were isolated from GCTB primary cell culture, and the role of p62 was evaluated using in vitro cell proliferation, migration and invasion assays. The results revealed that p62 mRNA and protein were overexpressed in tumor tissues. High p62 expression levels were significantly associated with the recurrence of GCTB (P=0.001). The patients in the high p62 expression group had shorter 5‑year recurrence‑free survival rates compared with the patients in the low p62 expression group (Pin vitro experiments revealed that p62 downregulation inhibited NS cell proliferation, invasion and migration, and promoted apoptosis. In conclusion, it was found that p62 overexpression is associated with the recurrence of GCTB via the promotion of NS cell proliferation. Therefore, p62 could be a novel prognostic indicator, and a potential therapeutic target for GCTB.