Objective To observe the curative effects of Jawei-Wendan decoction in treating acute exacerbation of COPD due to phlegm blocking in lung. Methods From July 2010 to June 2013, 66 enrolled cases were randomly divided into a treatment group and a control group with 33 in each. Aside from receiving anti-infection and symptomatic treatment as the control group did, the treatment group was also given orally Jawei-Wendan decoction. The symptoms, pulmonary function and the variety of the laboratory indexing were observed before and after the treatment. Results ①The treatment group attained better curative effects(P<0.05) and got shorter symptomatic relief time of cough, expectoration and breathless(The T-value is respectively 16.770, 8.635, 16.762, P<0.05). ②After 7-days treatment, the cure rate of the treatment group was 63.64%, and the control group was 39.39%. Their difference was conspicuous(P<0.05). The total effective rate in treatment group was 96.97%, which was 90.91% in control group. Their difference was inconspicuous(P>0.05).③The pulmonary function indices increased significantly in the treatment group[The values of FEV1、FVC and FEV1/FVC% are (1.41±0.46)L, (2.18±0.63)L and (66.41±6.33)% before treatment, which were (2.21± 0.55)L, (2.83±0.53)L and (77.37±7.64)%after treatment, P<0.01]. A comparison of the two groups showed significant differences regarding the pulmonary function indices(P<0.05). Conclusion Jawei-Wendan decoction had good curative effect in treating acute exacerbation of COPD due to phlegm blocking in lung.

Objective:To construct an expression system of lentivirus vector encoding epidermal growth factor receptor-specific chimeric antigen receptor (EGFR-CAR) and programmed cell death ligand-1 (PD-L1) antibody.Methods:Human PD-L1-Fc protein was used to immunize BALB/c mice. Cell-fusion and subcloning were performed to screen stable hybridoma strains with high secretion of PD-L1-specific antibodies, which were identified by both ELISA and Western blot. The activity of the antibodies in blocking the binding of programmed cell death-1 (PD-1) to PD-L1 was determined by fluorescence-activated cell sorting (FACS). Antibody affinity was analyzed by Fortebio Octet96. A single-chain variable fragment (scFv) was further constructed after antibody full-length sequencing and humanization using CDR grafting method. Meanwhile, the genes encoding the light and heavy chain variable regions (VL and VH) were cloned from a hybridoma secreting antibody against human EGFR by 5′ RACE technology to construct scFv gene. The expression of scFv was confirmed using pcDNA3.1 vector. EGFR-CAR containing CD137 intracellular function domain and PD-L1-scFv was ligated using 2A gene. The synthetic single molecule was cloned into pLVX-EF1a-IRES-ZsGreen1 lentivirus expression vector, and then transfected into 293T cells using Lenti-X Packaging Single Shots (VSV-G) to prepare infectious virus. Expression of CAR on cell surface and the soluble form of PD-L1-scFv in the supernatant of transfected 293V cells were detected by FACS and ELISA.Results:A PD-L1 antibody named 11E3 with high ligand-receptor blocking performance was obtained. The humanized antibody showed a stable affinity (2.67×10 -10 mol/L) after directly grafting the mouse CDRs (CDR1, CDR2 and CDR3) to human frameworks. EGFR-scFv was effectively expressed in a form of Fc-fusion. Secretory CAR (CTZ0431-1) and membrane CAR (CTZ0431-2) expression plasmids were constructed using lentivirus vector containing EGFR-CAR and PD-L1-scFv. The infection efficiency in 293V cells was around 10%. EGFR-scFv on the cell membranes and PD-L1-scfv in the culture supernatants were detected after 293V cells were infected with CTZ0431-1. EGFR-scFv and PD-L1-scfv were expressed on the cell membranes of 293V cells infected with CTZ0431-2. The expression rate of CAR in LV-CART46407-1-transfected activated T cells was 39.3%. Conclusions:The lentivirus vectors co-expressing EGFR-CAR with moderate binding affinity and PD-L1-scFv with high binding affinity were successful constructed, which provided an essential tool for investing EGFR- and PD-L1 double targeted CAR-T cell therapy against solid tumor.