Objective To investigate CBX2 and TIM3 in salivary adenoid cystic carcinoma(SACC)tissue and their relationship with clinical pathological features and prognosis.Methods 80 patients with SACC who underwent surgical treatment in the First Affiliated Hospital of Hebei North University from January 2016 to January 2020 were selected.Immunohistochemistry was used to measure CBX2 and TIM3 in tissues.The relationship between CBX2 and TIM3 in SACC tissue and prognosis was discussed though Kaplan-Meier method.The factors influencing the prognosis of SACC were discussed using multivariate Cox regression.Results The positive rates of CBX2 and TIM3 in SACC tissues were clearly higher than those in normal glandular tissues adjacent to cancer(χ2=11.237,8.229,P<0.05).The CBX2 and TIM3 were associated with nerve invasion and distant metastasis(P<0.05).After a 5-year follow-up,26 cases died and 54 cases survived,with an overall 5-year survival rate of 67.50%(54/80).The death group had higher positive rates of CBX2 and TIM3 in SACC tissues than the survival group(P<0.05).Patients with positive CBX2 and TIM3 in SACC tissues had clearly lower 5-year survival rate than patients with negative CBX2 and TIM3(Log Rank χ2=6.564,5.197,P<0.05).CBX2,TIM3 positivity,nerve invasion,and distant metastasis were risk factors affecting prognosis(P<0.05).Conclusion The positive expression of CBX2 and TIM3 in SACC tissues is closely related to the clinical pathological features and prognosis of patients.

Objective The mouse autologous tumor model H22 is more valuable for tumor immunological-related research.This paper aims to establish mouse hepatic tumor cell line(H22-luc2-tdT)that stably express the tan-dem-dimer tomato(tdTomato)and luciferase genes.Establish an in vivo imaging model of cell line derived trans-planted tumors.Methods Using transplanted H22 tumor tissue,primary culture and continuous passage in vitro were performed to establish a continuous cell line.Cell proliferation,chromosome analysis,organoid culture,tumorigenicity,HE and ICH of aFP,CK7,CK15 were performed to charaterize the cell line.Then the luc2-tdT plasmid was transfected into H22 cells of P22,flow cytometry and in vitro/in vivo imaging were employed to screen and verify fluorescence expression.Mycoplasma detection and species verification of the established cell lines were performed.Results The H22 cells had been continuously passaged over 50 times.The cells of passsge 22(P22)were transplanted subcutaneously and intraperitoneally into C57 and Kunming mice,with a 100%tumor formation.The HE morphology of subcutaneous transplanted tumor were consistent with the original tumor.CK+/AFP+proved that it was of liver cancer origin.The H22 cells were hypo-triploid with a modal number of 40-44 chromosomes and telocentromeres,verifing its mouse origin.The latent phase for in vitro growth of H22 lasted from d0 to d3,while the exponential phaes d3 to d5,and reach plateou at d6.Successful transfection of H22 cells with the luc2-tdT were observed with in vitro/in vivo 100%fluorescence positivity,thus named H22-luc2-tdT.The transplanted tumor tissue of H22 cells could be primarily cultured to form organoids.The detection of Mycoplasma was negative,and its mouse origin confirmed by PCR.Conclusions H22 and H22-luc2-tdT cell lines are established and characterized,which can be used for the establishment and application of in vitro and in vivo liver cancer research and metastatic cell tracking.These cell lines are deposited at and can obtain from the National Biomedical Cell Resource Center(http://www.cellresource.cn).

Objective To establish primary and simian virus 40(SV40)T antigen transformed human vascular en-dothelial cell models,and provide available resources for endothelial research.Methods Human umbilical vein endothelial cells(HUVEC),human umbilical artery endothelial cells(HUAEC),great saphenous vein endothelial cells(GSVEC)and endothelial cells form endometrium and liver tissue were isolated and cultured respectively.Then,the primary endothelial cells were transformed by lentivirus containing SV40 big T and small T antigens,and continuously subcultured in vitro.The expression of CD31 was detected by flow cytometry,species identification-and mycoplasma detection by PCR,and cell identity was identified by STR detection.The transformed ECs were checked for HLA types.Some of them were tested for RNA expression profile and infected by Cas9 lentivirus to es-tablish stable clones.Results Totally 187 cell lines of transformed HUVEC,1 of transformed HUAEC,5 of trans-formed GSVEC,1 of transformed endothelial cells from endometrium and 1 of transformed endothelial cells from liv-er tissue,and 9 monoclonal HUVEC cell lines stably expressing Cas9 protein were established.All the transformed umbilical endothelial cells were CD31 positive ranging from 20%-90%for 20 cases,while for the rest 168 cases the positive rate was more than 90%.RNA expression revealed stable activation of cell proliferation(cell cycle and DNA synthesis).Their species were identified as human origin.The STR results were consistent with those of the primary culture and unique,and there was no mycoplasma contamination.All these cells could be obtained with the sharing services of National Science and Technology Infrastructure,the National Biomedical Cell-line Resource cen-ters(NSTI-BMCR).Conclusions A series of primary and SV40 T antigen transformed human vascular endothelial cell models have been established,which provide a tool for the study of cardiovascular diseases,inflammation,tumors and immune-related diseases.

This review summarizes recent clinical progress in the field of lung cancer within the multidisciplinary team(MDT)diagnosis and treatment model across domestic and international studies.In the perioperative treatment of early-stage resectable non-small-cell lung cancer(NSCLC),the"sandwich"strategy combining preoperative neoadjuvant therapy with postoperative adjuvant therapy has demonstrat-ed significant survival benefits for patients.Notably,domestic toripalimab combined chemotherapy significantly increases the major patho-logic response(MPR)rate and event-free survival(EFS)rate of resectable stage Ⅲ NSCLC patients,emerging as a novel treatment regimen for perioperative NSCLC.For unresctable NSCLC patients,consolidative therapy combined with osimertinib,a third-generation epidermal growth factor receptor-tyrosine kinase inhibitor,following radical chemotherapy has significantly extended progression free survival,filling the gap in targeted therapy for stage Ⅲ lung cancer.Furthermore,the combination of immunotherapy and anti-angiogenic agents has ef-fectively improved the overall survival of advanced NSCLC patients.Significant progress has also been made in the field of antibody-drug conjugates,with agents such as sacituzumab and trastuzumab demonstrating substantial therapeutic efficacy.In the field of extensive-stage small-cell lung cancer(ES-SCLC),the bispecific antibody tarlatamab targeting Delta-like ligand 3(DLL3)and cluster of differentiation 3(CD3)has exhibited significant efficacy,and has received accelerated approval from U.S.Food and Drug Administration(FDA)as a sec-ond-line treatment for ES-SCLC.

This review summarizes recent clinical progress in the field of lung cancer within the multidisciplinary team(MDT)diagnosis and treatment model across domestic and international studies.In the perioperative treatment of early-stage resectable non-small-cell lung cancer(NSCLC),the"sandwich"strategy combining preoperative neoadjuvant therapy with postoperative adjuvant therapy has demonstrat-ed significant survival benefits for patients.Notably,domestic toripalimab combined chemotherapy significantly increases the major patho-logic response(MPR)rate and event-free survival(EFS)rate of resectable stage Ⅲ NSCLC patients,emerging as a novel treatment regimen for perioperative NSCLC.For unresctable NSCLC patients,consolidative therapy combined with osimertinib,a third-generation epidermal growth factor receptor-tyrosine kinase inhibitor,following radical chemotherapy has significantly extended progression free survival,filling the gap in targeted therapy for stage Ⅲ lung cancer.Furthermore,the combination of immunotherapy and anti-angiogenic agents has ef-fectively improved the overall survival of advanced NSCLC patients.Significant progress has also been made in the field of antibody-drug conjugates,with agents such as sacituzumab and trastuzumab demonstrating substantial therapeutic efficacy.In the field of extensive-stage small-cell lung cancer(ES-SCLC),the bispecific antibody tarlatamab targeting Delta-like ligand 3(DLL3)and cluster of differentiation 3(CD3)has exhibited significant efficacy,and has received accelerated approval from U.S.Food and Drug Administration(FDA)as a sec-ond-line treatment for ES-SCLC.

Objective To investigate CBX2 and TIM3 in salivary adenoid cystic carcinoma(SACC)tissue and their relationship with clinical pathological features and prognosis.Methods 80 patients with SACC who underwent surgical treatment in the First Affiliated Hospital of Hebei North University from January 2016 to January 2020 were selected.Immunohistochemistry was used to measure CBX2 and TIM3 in tissues.The relationship between CBX2 and TIM3 in SACC tissue and prognosis was discussed though Kaplan-Meier method.The factors influencing the prognosis of SACC were discussed using multivariate Cox regression.Results The positive rates of CBX2 and TIM3 in SACC tissues were clearly higher than those in normal glandular tissues adjacent to cancer(χ2=11.237,8.229,P<0.05).The CBX2 and TIM3 were associated with nerve invasion and distant metastasis(P<0.05).After a 5-year follow-up,26 cases died and 54 cases survived,with an overall 5-year survival rate of 67.50%(54/80).The death group had higher positive rates of CBX2 and TIM3 in SACC tissues than the survival group(P<0.05).Patients with positive CBX2 and TIM3 in SACC tissues had clearly lower 5-year survival rate than patients with negative CBX2 and TIM3(Log Rank χ2=6.564,5.197,P<0.05).CBX2,TIM3 positivity,nerve invasion,and distant metastasis were risk factors affecting prognosis(P<0.05).Conclusion The positive expression of CBX2 and TIM3 in SACC tissues is closely related to the clinical pathological features and prognosis of patients.

Traditional in vitro models have inevitable limitations and there are significant differences in assessing drug efficacy and side effects as compared to human trials.Organ-on-a-chip technology simulates human organs in a physiological environment and functional chip with a high fidelity physiological or pathophysiological level,offering great innovative prospects for drug development.This paper mainly introduces the research progress and application of organ chip from the perspective of various systems in vivo.At the same time,the limitations of the current devel-opment process of organ chip and the future development direction are proposed.

More and more in-depth tumor mechanism research and clinical precision treatment have put forward higher requirements for the technology for the establishment of tumor models.More and more tumor organoids have been applied.This model is able to reproduce the genomic,transcriptome,proteome and other omics features of pa-rental tumors without heterogeneity found in cell line models.Based on these characteristics,tumor organoids have gradually become a representative preclinical model,facilitating the translation of new research results and precision treatment of patients.This article summarizes the latest progress of tumor organoids on R&D of technology and appli-cation,focusing on the current methods of establishing tumor organoids,opportunities and challenges in clinical and research application.

Objective To establish breast cancer organoids for a long time and to characterize their molecular ex-pression and test their sensitivity to chemotherapy drugs.Methods Breast cancer specimens were digested with col-lagenase to release cancer cells for organoid culture.The organoids were characterized by morphology and ER,PR,HER-2,CK expression by technology of histoimmunofluorescence.Among them,three were tested for paclitaxel,doxorubicin and cisplatin sensitivity.Results A total of 44 cases of breast cancer organoids were established,all of which showed dense spherical-like growth and ER,PR,HER-2,CK,matching their clinical counterpart.Breast cancer organoids were sensitive to chemotherapy drugs paclitaxel,doxorubicin and cisplatin.Conclusions Organoid model,as a new in vitro may reproduce the pathophysiology features with heterogeneity.

Objective To establish human and mouse pancreatic cancer cell strains stably expressing Cas9 protein,green fluorescent protein,red fluorescent proteins,luciferase-tdTomato,and to validate the activity of luciferase and gene editing of Cas9 function for pancreatic cancer research using luciferase and CRISPR/Cas9 system.Methods In human pancreatic cancer cells(AsPC-1,CFPAC-1,HPAC,BxPC-3,HS 766T,MIA PaCa-2,PANC-1,and SW 1990),and mouse pancreatic cancer cell(Pan02),the cells were infected with Cas9-expressing plas-mid pLv-EF1α-Cas9m1.1-Puro,and single-cell clones were selected for culture and expansion.After extracting the total protein,Western blot verified the expression level of Cas9;Infected with fluorescent protein expression plasmids pLv-EF1α-EGFP,pLv-EF1α-mCherry,pLv-EF1α-tdTomato,pLv-EF1α-Luc2-tdT,and selected single cell clones stably expressing fluorescent proteins were cultured and amplified under fluorescence microscope.Cas9 stable expression cell line was selected to be infected with pLv-EF1α-Luc2-tdT,and the mono-clonal culture of stable expression of fluorescent proteins was selected for expansion under fluorescence micro-scope.One of the cell lines were selected to be infected with Lv-EF1a-mCherry,and the mCherry-positive cells were sorted out by flow cytometry,and then the guide RNA of mCherry gene was then infected by lentivirus to tar-get the mCherry gene,and after cell expansion,mCherry knockdown was detected by fluorescence microscope observation and flow cytometry;5 BALB/c Nude mice were subcutaneously inoculated with MIA PaCa-2-Luc2-tdT cells(1.0×107/cells each),and imaged in vivo after 36 days.Results 48 human pancreatic cancer cell strains with stable Cas9 expression were screened(including 23 cells expressing Cas9m1.1,25 cells expressing Cas9m1.1-Luc2-tdT),33 pancreatic cancer cell strains with stable expression of fluorescent proteins were screened(8 cells expressing EGFP,7 expressing mCherry,and 9 each expressing Luc2-tdT and tdTomato).Cells expressing mCherry and Cas9 were infected with mCherry gRNA and mCherry was knocked down.In vivo imaging showed that both bioluminescence and fluorescence luminescence were present in MIA PaCa-2 cells ex-pressing Luc2-tdT.Conclusions 33 pancreatic cancer cell strains with stable expression of fluorescent proteins are successfully established,in which the Luc2-tdT-expressing cell strains have luciferase activity;48 pancreatic cancer cell strains with stable expression of Cas9 are successfully established,and the Cas9 protein has gene edi-ting activity,gene editing activity varies depending on the original cell strains.