Cholangiocarcinoma (CCA) has emerged as an intractable cancer with scanty therapeutic regimens. The aberrant activation of Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) are reported to be common in CCA patients. However, the underpinning mechanism remains poorly understood. Deubiquitinase (DUB) is regarded as a main orchestrator in maintaining protein homeostasis. Here, we identified Josephin domain-containing protein 2 (JOSD2) as an essential DUB of YAP/TAZ that sustained the protein level through cleavage of polyubiquitin chains in a deubiquitinase activity-dependent manner. The depletion of JOSD2 promoted YAP/TAZ proteasomal degradation and significantly impeded CCA proliferation

Objective:To compare the histologic features of immune-mediated hepatitis (IMH) due to immune checkpoint inhibitors (ICIs) monotherapy and combined ICIs anti-angiogenesis tyrosine kinases (TKIs) targeted therapy.Methods:Twenty-one IMH patients who had liver biopsy during ICIs treatment in Zhongshan Hospital of Fudan University from 2015 to 2019 were included. Among them, ten were treated with ICIs monotherapy, and 11 were treated with combined ICIs and anti-angiogenesis targeted therapy. The histologic features of IMH were assessed by HE staining and PD-L1/2 was evaluated by immunohistochemical staining.Results:Patients treated with monotherapy ICIs presented with different levels of lobular hepatitis and portal inflammation. Besides, there were also cholangitis, endothelialitis, Kupffer cells activation and peliosisi hepatitis. Eight cases (8/10) showed mild and two cases (2/10) showed moderate hepatic injury. As for patients receiving combined ICIs and TKIs therapy, the extent of IMH was more severe, with four cases (4/11) showing moderate-severe liver injury, with confluent or bridging necrosis, portal inflammation, cholangitis, interface hepatitis. Among these, one patient developed acute severe hepatitis with massive hepatocyte necrosis and died of multisystem dysfunction. In those cases with severe liver injury, many CD8 positive lymphocytes aggregated in the portal area and hepatic sinusoid, and PD-L1 was expressed in many endothelial cells. There were both 2 cases of death in ICIs monotherapy and combination therapy group. Among the latter group, 1 patient developed acute severe hepatitis with massive hepatocyte necrosis and died of multisystem dysfunction.Conclusion:Compared with ICIs monotherapy, combined ICIs and anti-angiogenesis targeted TKIs therapy may cause overlapping hepatic injury, leading to severe IMH.

Objective:To analyze the different expressions of miRNAs in peripheral blood of rheumatoid arthritis (RA) patients and the osteoclasts in rats,and to verify the key miRNAs in RA.Methods: The miRNAs expressions in monocyte and the co-cultured osteoclasts,peripheral blood of rheumatoid arthritis (RA) patients and normal were detected by miRCURYTM LNA Array.Real time PCR was applied to verify the reliability of miRNA array.The bioinformatics software and database were applied to predict and analyze target genes.Results: miRNA array results showed that 189 miRNAs changed in RA patients compared with the normal;211 miRNAs were changed in osteoclasts group compared with monocytes group.The expressions of ten miRNAs were all abnormal in RA patients and osteoclasts in rats.The results of Real time PCR were consistent with the array.Results of bioinformatics analysis showed that the target gene of miRNA significantly enriched in signaling pathways such as VEGF,MAPK signaling pathways.Conclusion: Some miRNAs had similar abnormal expressions in different species,and these miRNAs may influence the differentiation of the osteoclasts through regulating the related signal passways.

Objective: To explore the mechanism of ibrutinib on drug resistance diffuse large B-cell lymphoma (DLBCL) cells. Methods: DLBCL cell line was cultured with mesenchymal stem cells (MSC) , and DLBCL cells which migrated and adhered to MSC under microscope was counted. The secretion of CXCL12 by MSC were measured by ELISA. The expression of CXCR4 on DLBCL cells were measured by flow cytometry, HBL-1 cells were transfected with a CXCR4-lentivector. An Annexin Ⅴ-binding assay was used to detect the induction of apoptosis. Clonogenic growth of DLBCL cells was evaluated on MethoCult media. Ibrutinib was injected into NOD/SCID mice, tumor growth was assessed via caliper measurements every 3 days. Results: MSC promoted migration and adhesion of DLBCL cells to MSC. Ibrutinib inhibited migration and adhesion of DLBCL cells to MSC in a dose-dependent manner (P<0.05) . CXCL12 secreted by MSC and CXCR4 expressed on DLBCL cells could induce each other, which upgraded the levels of secretion and expression. Ibrutinib could inhibit the secretion of CXCL12 (SUDHL10: 660 pg/ml vs 1 400 pg/ml, P=0.004; HBL-1: 720 pg/ml vs 1 490 pg/ml, P=0.018; DLBCL:850 pg/ml vs 1 450 pg/ml, P=0.004) and expression of CXCR4 (P<0.05) . When co-cultured with MSC, the ratio of HBL-1 cells apoptosis in the group of control, mitoxantrone, ibrutinib, mitoxantrone+ibrutinib were respectively 15.1%, 17.5%, 23.5%, 58.7%. After transfected with a CXCR4-lentivector and overexpressed CXCR4, the ratios of HBL-1 cells apoptosis were 14.2%, 16.1%, 22.5%, 38.3% respectively. The ratio of DLBCL cells apoptosis induced by mitoxantrone was lower when co-cultured with MSC (P<0.05) . But with the addition of ibrutinib, the ratio of apoptosis was increaed and it was similar to cultivation without MSC, which suggested ibrutinib could inhibit drug-resistance induced by MSC. But after transfected with a CXCR4-lentivector, the overexpression of CXCR4 was detected and the ratio of apoptosis was significantly lower when co-cultured with MSC which demonstrated that ibrutinib inhibited drug-resistance by inhibiting the expression of CXCR4. MSC enhanced lymphoma clonogenicity in vitro and lymphoma cell growth in vivo. The number of colonies of control, MSC, Ibrutinib, MSC+Ibrutinib were 113±5, 205±4, 62±9, 123±3 (2.5×10³/well, ±s) , respectively. The tumor volume of NOD/SCID mice were respectively 6 500, 17 000, 4 000, 10 000 mm³. Ibrutinib inhibited lymphoma clonogenicity in vitro and lymphoma cell growth in vitro. Conclusion Ibrutinib targeted the CXCL12/CXCR4 axis, inhibited the expression of CXCR4 and inhibited MSC-mediated drug resistance. Ibrutinib also inhibited lymphoma clonogenicity in vitro and lymphoma cell growth in vivo. These results provided a scientific rationality for relapsed/refractory DLBCL treatment with ibrutinib.

Objective To evaluate the impacts of slice thickness and registration frame range on the accuracy of cone-beam computed tomography ( CBCT) image-guided head and neck ( HN) radiotherapy, and to provide a basis for positioning correction in image-guided radiotherapy.Methods A planned CT scan was performed for an anthropomorphic HN phantom with slice thickness of 1 mm and 3 mm and simulated positioning errors in x, y, and z directions on the accelerator.CBCT scan and reconstruction were performed with slice thickness of 1 mm and 3 mm.Two different registration frame ranges were used ( range 1:from C7 to superior orbit;range 2:from C7 to calvaria ) .Automatic bony registration was performed for CBCT and planned CT images with slice thickness of 1 mm and 3 mm.The registration accuracy was evaluated.Results For range 1, the registration errors in x, y, and z directions with a slice thickness of 1 mm were significantly lower than those with a slice thickness of 3 mm (0.5±0.2 mm vs.-0.7±0.2 mm, P=0.00;0.5±0.3 mm vs. 1.0±0.3 mm, P=0.00;-0.1±0.5 mm vs.1.5±0.5 mm, P=0.00).For range 2, the registration errors in x, y, and z directions with a slice thickness of 3 mm were-0.4±0.2 mm, 0.5±0.2 mm, and 0.7±0.4 mm, respectively.Conclusions Engagement of calvaria in registration range can substantially enhance the registration accuracy in CBCT or CT images for HN.The registration error with slice thickness of 1 mm can be controlled within 1 mm.

Objective:To detect the inhibitory effects of CAL-101, a selective inhibitor of phosphoinostitide-3'-kinase delta (PI3Kδ), on Burkitt's lymphoma cell line Raji and diffused large B-cell lymphoma cell line SUDHL-10 and elucidate its relative mechanism. Methods:Raji and SUDHL-10 cells were treated with various concentrations of CAL-101. Methyl thiazolyl tetrazolium (MTT) assay was performed to determine the inhibitory effect of CAL-101 on lymphoma cells, and cell apoptosis was measured by Annexin V/PI and DAPI staining. Migration assays were performed with transwell to detect the migration of lymphoma cells derived from the stromal cell line HK. Western blot was used to detect the phosphorylation status of the ERK pathway. MTT and CalcuSyn software analyses were preformed to detect whether or not combining CAL-101 with bortezomib induces synergistic cytoxicity. Results:CAL-101 at con-centrations of 5, 10, 15, and 20μmol/L inhibited cell proliferation in a dose-dependent manner. The proliferation rates of the Raji cells treated with 5, 10, 15, and 20μmol/L for 48 h were 29.17%± 1.23%, 38.15%± 1.51%, 46.46%± 1.78%, and 55.8%± 2.01%, respec-tively, which were significantly higher (P<0.05) than that of the control group (1.15% ± 0.02%). Similar results were found in the SUDHL-10 cells after treatment with CAL-101 (P<0.05). CAL-101 also exerted an apoptotic effect on the lymphoma cells. The apop-totic rates of the Raji cells treated with CAL-101 for 21 h were 22.69%± 3.83%and 49.96%± 7.36%, respectively, which were signifi-cantly higher (P<0.05) than that of the control group (5.23%± 2.04%). Similar results were found in the SUDHL-10 cells (P<0.05). Treatment with 5 and 10 μmol/L CAL-101 dose-dependently inhibited the migration activity of lymphoma cells to stromal cells (P<0.05). Western blot analysis showed that the expression level of ERK phosphorylation protein was significantly downregulated in the cells treated with CAL-101. A synergistic effect between CAL-101 and bortezomib was verified. That is, these two drugs can signifi-cantly inhibit the proliferation of lymphoma cells with CI values less than 1. Conclusion:The PI3Kδ-specific inhibitor CAL-101 sup-pressed the proliferation of Raji and SUDHL-10 cells, induced apoptosis, and inhibited stromal cell-derived migration. This inhibitory effect may be induced by blocking the ERK pathway. Overall, our study indicated that CAL-101 is a novel and potential agent in the therapeutic strategy against aggressive B-cell lymphoma.

Objective To evaluate the accuracy of image registration based on bony structure (RBS) and grey-scale (RGS) in positioning correction of radiation treatment,and their reliability in clinical application.Methods Setup errors of anthropomorphic phantom (chest& abdomen,head& neck) were simulated with x-,y-,z-directions.CBCT images were acquired for each simulation and registered with planning CT.using bony structure and grey-scale registration separately.Geometry accuracy of all registration were then obtained and analyzed.Results The errors of RBS and RGS in x-,y-,z-directions were (-0.65 ±0.22) mm and (-0.70±0.17) mm (P=0.00),(1.02 ±0.27) mm and (0.90±0.20) mm (P =0.00),(1.46 ± 0.53) mm and (1.47 ± 0.47) mm (P =0.54) for head& neck positioning; with (0.82±0.33) mm and (0.79±0.18) mm (P=0.03),(2.45±1.17) mm and (1.61 ±0.84) mm (P =0.00),(1.44 ± 3.25) mm and (0.19 ± 1.11) mm (P =0.00) for chest& abdomen positioning.Conclusions RGS is more accurate and stable than RBS.The accuracy of image registration is a little better for head& neck than that for chest& abdomen.The algorithms of image registration used in clinical application needs to be tested independently and the systematic error needs to be corrected before applying in different treatment techniques according to their accuracy requirement.

ObjectiveTo evaluate the local failure and the impact on survival by prospectively comparing involved field radiotherapy (IFRT) and elective nodal irradiation (ENI) in combination with concurrent chemotherapy for locally advanced non-small cell lung cancer ( LA-NSCLC ).Methods LANSCLC patients were treated with 2 cycles of carboplatin ( AUC =5 - 6,d1 ) combined with paclitaxel ( 175mg/m2 ),followed assessment without distant metastasis,then randomized into IFRT (45 patients) or ENI (54 patients) arm.IFRT included primary tumor,ipsilateral hilar and positive mediastinal lymph nodes;ENI included the primary lesion,ipsilateral hilar,hilateral mediastinal lymph node drainage and bilateral supraclavicular area.The prescription dose was given as high as possible with V20 ≤35％ and spinal cord dose ≤50 Gy,combined weekly paclitaxel 40 mg/m2 concurrent chemotherapy.The Kaplan-Meier method was used to estimate survival data and the log-rank method was used to test distribution of survival time between arms.ResultsThe follow-up rate was 99％.49,29 and 17 patients were followed-up for 1-,2-and 3-year,respectively.More patients from group IFRT received ＞60 Gy than ENI (49％ vs.26％,x2 =5.59,P =0.018 ).The local failure rates were 29％ and 36％,respectively ( x2 =0.46,P =0.497 ).The 1-,2-and 3-year local tumor progression-free survival rates were 76％,69％,65％ and 80％,53％,49％ ( x2 =0.74,P =0.389),respectively; the 1-,3-and 5-year overall survival rates were 80％,41％,33％ and 69％,32％,13％ (x2 =3.97,P =0.046),respectively.There were no significant differences in acute and late toxicities between the arms ( x2 =3.910 - 0.155,P =0.142 - 0.925 ).ConclusionsIFRT improved radiation dose and survival rate and did not increase the failure of elective lymph node region compared with ENI.The toxicities were no differences between IFRT and ENL Further investigation with big size sample is warranted.

Objective To investigate the expressions of Sema4D and HER-2 in breast cancers and their relationship with microlymphtic vessel density (MLVD).Methods MLVD and expressions of Sema4D and HER-2 were detected in 110 cases of breast cancer tissues by immunohistochemistry.The relationship of Sema4D and HER-2 expressions with clinicopathologic parameters was analyzed.Results The positive expression rate of Sema4D and HER-2 was 71.82％ (79/110) and 33.64％ (37/110),respectively,with a statistically significant differences compared to those in the control group ( P ＜0.01 ).The positive expression rates of Sema4D and MLVD in cases with lymphatic metastasis were higher than that without lymphatic metastasis ( P ＜ 0.01 ).The expressions of Sema4D and HER-2 were closely correlated with the histological grade,TNM stage,lymph node metastasis,and expression of ER of breast cancers ( P ＜0.05),but were not related to tumor size and expression of PR (P ＞ 0.05 ).On univariate analysis,the disease-free survival rate of patients with Sema4D positive expression was better than those with its negative expression ( P ＜ 0.05).Sema4D expression was positively correlated with the HER-2 expression ( r =0.535,P ＜ 0.01 ).Conclusions Sema4D may play important roles in the carcinogenesis and development of a breast cancer.

Ninety six female patients with chronic renal failure were randomly allocated into combination group (n =48) and control group (n =48).In combination group patients received both kidney transplantation and hematopoietic stem cell infusion,in control group patients underwent kidney transplantation only.The results showed that chronic rejection in the combination group was lower than that in the control group [2％(1/48)vs.17％ (8/48),P＜0.05)].The 1-,3-,5-and 10 y-survival rates of kidney in the combination group were 98％ (47/48),94％ (45/48),83％ (34/41) and 9/17,respectively,those in control group were 98％ (47/48),90％ (43/48),76％ (31/41) and 7/17,respectively.Infusion of donor hematopoietic stem cells can augment chimerism in early postoperative period and significantly reduce the rate of graft rejection,which is beneficial for the quality of life of the recipients.