[摘 要] 目的：探究肺癌磷酸甘油酸变位酶1（PGAM1）对肺癌LLC细胞增殖和迁移的调控作用，以及PGAM1对肿瘤微环境中CD8+ T细胞功能和浸润的影响。方法：将shPGAM1及shNC慢病毒感染LLC细胞，筛选稳定细胞，分别命名为shPGAM1组和NC组，WB和qPCR法检测两组细胞中PGAM1蛋白和mRNA的表达水平。CCK-8法和实时细胞分析仪检测敲低PGAM1对LLC细胞增殖和迁移的影响；流式细胞术检测与两组细胞分别体外共培养后CD8+ T细胞功能性受体（TIM-3、PD-1、GrzmB、Ki67）表达的变化。构建小鼠肺癌LLC细胞皮下移植瘤模型，监测PGAM1表达对肿瘤生长的影响，通过流式细胞术检测敲低PGAM1对肿瘤微环境中CD8+ T细胞浸润的影响。结果：shPGAM1组细胞中PGAM1蛋白和mRNA表达水平较NC组均显著降低。敲低PGAM1可降低LLC细胞的增殖和迁移能力（Ρ < 0.000 1或Ρ < 0.05）。与shPGAM1组细胞共培养后，CD8+ T细胞的耗竭标志物（TIM-3、PD-1）表达均降低（Ρ < 0.000 1或Ρ < 0.01）。敲低PGAM1后肿瘤生长受到显著抑制，且肿瘤组织中CD8+ T细胞浸润增加（Ρ <0.000 1）。结论：靶向敲低PGAM1可抑制肿瘤细胞增殖、迁移，降低CD8⁺ T细胞耗竭标志物的表达及促进肿瘤内T细胞浸润，双重调控肿瘤生长与抗肿瘤免疫。

Malignant tumor is one of the main diseases that seriously threaten human health and life.The incidence and mortality have in-creased year by year.In China,malignant tumors have become the leading cause of death among residents.For a long time,surgery,radio-therapy and chemotherapy have become the three conventional treatment methods of tumor treatment,but many tumors cannot achieve effective treatment.With the changes in the concept of tumor treatment,immunotherapy has become the fourth model of comprehensive tumor treatment and has received increasing attention.On the basis of existing evidence-based medical results,the China Anti-Cancer Asso-ciation(CACA)combines domestic and foreign guidelines and consensus to formulate a guide for the immunotherapy in malignant tumors,hoping to help physicians engaged in clinical immunotherapy.

[摘 要] 目的：探讨结直肠癌（CRC）组织中磷酸甘油酸变位酶1（PGAM1）的表达及其与患者预后的关系，研究PGAM1对CRC细胞增殖、迁移和侵袭的影响。方法：选择2003年3月至2008年11月间在天津医科大学肿瘤医院手术切除的30例CRC患者的肿瘤组织标本及临床资料，采用免疫组织化学染色法检测CRC组织中PGAM1蛋白的表达，分析PGAM1表达与患者临床病理特征的关系，Kaplan-Meier生存分析法比较PGAM1高表达与低表达患者的OS、PFS来评价PGAM1表达与患者预后的关系。利用RNA干扰技术分别将si-PGAM1及si-NC质粒转染至HCT-116和SW480细胞，WB法检测转染细胞中PGAM1蛋白的表达水平，CCK-8、Transwell实验分别检测敲低PGAM1对CRC细胞增殖、迁移和侵袭的影响。结果：30例CRC组织中PGAM1阳性染色定位于CRC细胞的细胞质，其中33.3%（10/30例）呈高表达。虽然PGAM1高表达与CRC患者年龄、性别、组织学类型、肿瘤大小、淋巴结转移、远处转移及临床TNM分期无关（均P>0.05），但是PGAM1高表达与低表达患者相比其OS、PFS显著缩短。在CRC细胞中敲低PGAM1后，细胞的增殖、迁移和侵袭能力均显著降低（均P<0.05）。结论：CRC组织中PGAM1呈高表达，PGAM1高表达的患者预后较差；敲低PGAM1后细胞的增殖、迁移及侵袭能力均显著降低，提示PGAM1可能是CRC患者预后的生物标志物。

BACKGROUND: Small cell lung cancer (SCLC) with high c-Myc expression is prone to relapse and metastasis, leading to extremely low survival rate. Cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitor Abemaciclib plays a key role in the treatment of tumors, but the effects and mechanisms on SCLC remain unclear. This study was to analyze the effect and molecular mechanism of Abemaciclib in inhibiting proliferation, migration and invasion of SCLC with high c-Myc expression, with a view to expanding a new direction for reducing the recurrence and metastasis. METHODS: Proteins interacting with CDK4/6 were predicted using the STRING database. The expressions of CDK4/6 and c-Myc in 31 cases of SCLC cancer tissues and paired adjacent normal tissues were analyzed by immunohistochemistry. The effects of Abemaciclib on the proliferation, invasion and migration of SCLC were detected by CCK-8, colony formation assay, Transwell and migration assay. Western blot was used to detect the expressions of CDK4/6 and related transcription factors. Flow cytometry was used to analyze the effects of Abemaciclib on the cell cycle and checkpoint of SCLC. RESULTS: The expression of CDK4/6 was associated with c-Myc by STRING protein interaction network. c-Myc can directly modalize achaete-scute complex homolog 1 (ASCL1), neuronal differentiation 1 (NEUROD1) and Yes-associated protein 1 (YAP1). Moreover, CDK4 and c-Myc regulate the expression of programmed cell death ligand 1 (PD-L1). Immunohistochemistry showed that the expressions of CDK4/6 and c-Myc in cancer tissues were higher than those in adjacent tissues(P<0.0001). CCK-8, colony formation assay, Transwell and migration assay verified that Abemaciclib could effectively inhibit the proliferation, invasion and migration of SBC-2 and H446OE(P<0.0001). Western blot analysis further showed that Abemaciclib not only inhibited CDK4 (P<0.05) and CDK6 (P<0.05), but also affected c-Myc (P<0.05), ASCL1 (P<0.05), NEUROD1 (P<0.05) and YAP1 (P<0.05), which are related to SCLC invasion and metastasis. Flow cytometry showed that Abemaciclib not only inhibited the cell cycle progression of SCLC cells (P<0.0001), but also significantly increased PD-L1 expression on SBC-2 (P<0.01) and H446OE (P<0.001). CONCLUSIONS Abemaciclib significantly inhibits the proliferation, invasion, migration and cell cycle progression of SCLC by inhibiting the expressions of CDK4/6, c-Myc, ASCL1, YAP1 and NEUROD1. Abemaciclib can also increase the expression of PD-L1 in SCLC.
Humans ; Small Cell Lung Carcinoma ; B7-H1 Antigen ; Sincalide ; Lung Neoplasms ; Neoplasm Recurrence, Local ; Transcription Factors ; Adaptor Proteins, Signal Transducing ; Cell Proliferation

[Abstract] Objective: To investigate the relationship between 18F-FDG PET/CT metabolic indicators and expression of immunocyte markers in lung adenocarcinoma patients, and to explore its significance in treatment and prognosis prediction for lung adenocarcinoma patients.Methods:Theclinicaldataof85lungadenocarcinomapatients,whoadmittedtoTianjinMedicalUniversityCancerInstituteand Hospital and underwent PET/CT examination from April 2008 to August 2014, were retrospectively analyzed. The expression levels of CD3, CD8, CD68, CD163, CD11c, Foxp3, PD-1 and PD-L1 were determined by immunohistochemistry. Correlations among immune markers (CD68+TAM), PET/CT metabolic parameters (SUVmax, SUVpeak and SUVmean) and tumor metabolic indicators (MTV , TLG) were analyzed using Pearson correlation analysis.The relationships between tumor metabolism, immune indicators and patients’survival outcomes were analyzed using the Kaplan-Meier method. Results: There was a remarkably negative correlation between SUVmax, SUVpeak, SUVmean and expression level of CD68+TAMs(r=-0.253,-0.265,-0.263,allP<0.05)butpositivecorrelationwithPD-1+TILs (r=0.427, 0.402, 0.395, all P<0.01) in lung adenocarcinoma patients. MTV and TLG were positively associated with Foxp3+ Tregs and PD-1+ TILs (r=0.313, 0.307, 0.29, 0.407, all P<0.01). Kaplan-Meier survival analysis showed that SUVmax, SUVmean, CD11c+DCs, PD-L1+ cells and TLG were all significantly associated with patients’prognosis (PFS or OS) (all P<0.05). Conclusion: Metabolism of tumor primary lesions is significantly correlated with tumor infiltrating immunocytes, and some of these indicators were associatedwithpatients’prognosis, suggesting that tumor metabolism and microenvironment immune status reflected by 18F-FDG PET/CTindicatorsmay have important reference value fortheimmunotherapyandprognosispredictionoflungadenocarcinoma patients.

With the progress of gene detection technology and the speed-up in new drug development, biological target therapy has fully covered the first-line treatment of advanced NSCLC. Immunotherapy has significantly improved the survival of advanced NSCLC patients with negative driven genes, and the median OS reaches about 2 years (15.6-30 months). EGFR is the most common driven gene. According to different EGFR mutation subtypes (L858R or 19del), different treatment mode (EGFR-TKI single drug, TKI combined with anti-vascular drugs and TKI combined with chemotherapy) is selected as the first-line treatment, which has become a consensus. Depending on the data of median PFS, the treatment efficacy against rare targets is more prominent, which has exceeded the efficacy of standard chemotherap:ALK (alectinib, PFS=34.8 months), ROS1 (ceritinib, PFS=19.3 months), RET (selpercatinib, PFS=18.4 months), BRAF (dabrafenib plus trametinib, PFS=14.6 months), NTRK (larotrectinib, PFS≥12 months) and MET (savolitinib, PFS=9.7 months). In conclusion, the first-line treatment of advanced NSCLC has entered the era of“precision-targeted treatment”based on different molecular typing, and it has become a consensus that high-throughput sequencing is required for newly diagnosed patients.

PD-1/PD-L1 (programmed cell death 1/programmed cell death 1 ligand 1) and CTLA-4 (cytotoxic T lymphocyte antigen 4) are currently approved major immune checkpoints. Immune checkpoint inhibitors against them are novel monoclonal antibodies that perform well in a variety of malignancies such as melanoma, renal cell carcinoma, non-small-cell lung cancer, urothelial carcinoma and Hodgkin′s lymphoma. However, with the increasing use of immune checkpoint inhibitors, immune-related adverse events cannot be ignored. The incidence of gastrointestinal toxicity is second only to skin toxicity. In this review, we focused on the mechanisms of these immune checkpoint inhibitors and the characteristics of gastrointestinal toxicity induced by them, and also discussed the clinical management strategies.

Chimeric antigen receptor T-cell (CAR-T) is one of the effective methods for treatment of lymphoma. The way to improve the efficacy and control the reverse reactions still needs to be explored further. Several clinical trials have indicated CAR-T could have favorable effects on the B-cell lymphoma patients with controllable reverse reactions. However, antigen loss is a major factor for the acquired resistance to CD19 CAR-T therapy. Other clinical researches, including CD22 for treatment of B-cell lymphoma and CD30 for Hodgkin lymphoma, have increased the efficacy of CAR-T. Moreover, lots of trials have suggested that the patients who received cyclophosphamide or bendamustine plus fludarabine lymphodepletion can get a high effective rate.

PURPOSE: The treatment of triple-negative breast cancer (TNBC) remains challenging, due to the absence of estrogen, progesterone, and human epidermal growth factor receptors. This study was designed to evaluate the efficiency and safety of cytokine-induced killer (CIK) cell immunotherapy, following regular chemotherapy, for patients with TNBC. METHODS: A total of 340 patients with postmastectomy TNBC, from January 1, 2010 to June 30, 2014, were included in this retrospective study. Seventy-seven patients received CIK cell immunotherapy, following regular chemotherapy (arm 1), and 263 patients received regular chemotherapy alone (arm 2). The primary aim was overall survival (OS) and disease-free survival (DFS), and the treatment responses and adverse events were also evaluated. RESULTS: The 5-year DFS and OS rates in arm 1 were 77.9% and 94.3%, compared with 69.8% and 85.6% in arm 2, respectively (p=0.159 and p=0.035, respectively). This clearly shows that there was no statistical difference in the 5-year DFS between the two groups. Multivariate analyses of arm 1 indicated that a Karnofsky performance score (KPS) ≥90 and stage I/IIA disease were significantly associated with a prolonged DFS period (hazard ratio [HR], 0.25; 95% confidence interval [CI], 0.09–0.74; p=0.012; and HR 0.21; 95% CI, 0.06–0.82; p=0.024, respectively), but a KPS ≥90 and stage I/IIA disease were not independent prognostic factors for OS. Toxicity was mild in patients who received the CIK therapy. CONCLUSION: The data suggested that CIK cell immunotherapy improved the efficiency of regular chemotherapy in patients with TNBC, and the side effects of CIK cell immunotherapy were mild.
Arm ; Cytokine-Induced Killer Cells* ; Disease-Free Survival ; Drug Therapy* ; Estrogens ; Humans ; Immunotherapy ; Multivariate Analysis ; Progesterone ; Prognosis ; Receptor, Epidermal Growth Factor ; Retrospective Studies ; Triple Negative Breast Neoplasms*

Objective: To investigate the effect of myeloid-derived suppressor cells (MDSCs) from mice bearing breast cancer on the function of normal B cells. Methods:ABABL/c mouse 4T1 breast cancer model was established. The spleen MDSCs of tumor-bearing mouse and normal mouse spleen B cells were sorted by magnetic beads, and the sorted MDSCs and B cells were co-incubated. Flow cytometry was used to test the effect of MDSCs on the expressions of B cell surface molecules, including PD-1, PD-L1, CTLA-4, CCR6, CD62L and MHCⅡ; ELISA assay was used to detect the secretion of IgA, IgM and IgG by B cells; BrdU kit was used to detect B cell proliferation; andAnnexin V/PI staining was used to detect B cell apoptosis. B cells in the co-culture system were again sorted by magnetic beads and were then co-cultured with T cells; BrdU kit was used to detect T cell proliferation, and Annexin V/PI was used to detect T cell apoptosis. Results: Compared with B cell control group, the expression of PD-L1 on B cells in B+MDSC group was increased (P<0.01), while the expressions of PD-1, CTLA-4, CCR6, CD62L and MHC Ⅱ were all decreased (all P<0.01); The IgA, IgM and IgG secreted by B cells were significantly increased (all P<0.01); the proliferation of B cells was increased (P<0.01) and the apoptosis was decreased (P<0.01). Compared with the T cell control group, the proliferation of T cells in the B+MDSC (1:5) +T group was significantly reduced (P<0.01); however, there was no significant difference in T cell apoptosis. Conclusion: MDSCs from breast cancer bearing mice promotes B cell proliferation and inhibits B cell apoptosis, and the MDSC-induced B cells can inhibit T cell proliferation.