Phase 1 clinical trial to assess safety and efficacy of NY-ESO-1-specific TCR T cells in HLA-A∗02:01 patients with advanced soft tissue sarcoma.

New York esophageal squamous cell carcinoma-1 (NY-ESO-1)-specific T cell receptor (TCR) T cell therapy is effective in tumors with NY-ESO-1 expression, but a safe and effective TCR-T cell therapeutic protocol remains to be improved. Here, we report a phase 1 investigational new drug clinical trial with TCR affinity-enhanced specific T cell therapy (TAEST16001) for targeting NY-ESO-1. Enrolled patients receive TAEST16001 cell infusion after dose-reduced lymphodepletion with cyclophosphamide (15 mg/kg/day × 3 days) combined with fludarabine (20 mg/m2/day × 3 days), and the TCR-T cells are maintained with low doses of interleukin-2 injection post-adoptive transfer. Analysis of 12 patients treated with the regimen demonstrates no treatment-related serious adverse events. The overall response rate is 41.7%. The median progression-free survival is 7.2 months, and the median duration of response is 13.1 months. The protocol of TAEST16001 cells delivers a safe and highly effective treatment for patients with advanced soft tissue sarcoma (ClinicalTrials.gov: NCT04318964).

Causal relationship between ischemic stroke and its subtypes and frozen shoulder: a two-sample Mendelian randomization analysis.

Background: Previous epidemiological and other studies have shown an association between ischemic stroke (IS) and frozen shoulder (FS). However, the causal relationship between them remains unclear. Therefore, the present study aimed to investigate the causal relationship between IS and FS using a two-sample Mendelian randomization method. Methods: Our research was divided into two stages: discovery and replication. The data were extracted from publicly available genome-wide association studies (GWAS). We selected a large sample of IS (n = 440, 328) and its subtypes (large-artery atherosclerotic stroke (LAS), cardioembolic stroke (CES), and stroke caused by small-vessel disease (SVS) and lacunar stroke (n = 254, 959) as exposure data. Additionally, we selected a large sample of FS as outcome data (n = 451, 099). Inverse variance weighting (IVW) was applied as the primary analysis method. The weighted median, MR-Egger, simple model, and weighted model were used as complementary analysis methods to assess causal effects. Moreover, heterogeneity was analyzed using Cochran's Q-test with IVW and MR-Egger. The MR-Egger intercept and MR-PRESSO analysis methods were used for pleiotropy testing. The stability of the results was also assessed using a leave-one-out analysis. Results: In the discovery stage, the IVW approach revealed an odds ratio (OR) of 1.207 with a 95% confidence interval (CI) of 1.027-1.417 and a P-value of 0.022. This suggests a causal association between IS levels and an increased risk of FS. In the subtype studies of IS, the findings were negative. However, during the replication stage, a significant causal link was found between selected lacunar strokes and FS with an OR of 1.252, a 95% CI of 1.105-1.419, and a P-value of 0.0004. All studies had no pleiotropy or heterogeneity, and the findings were robust. Conclusions: Our study confirmed the causal relationship between any IS level and increased risk of FS. Furthermore, the same results were obtained in the replication stage with lacunar stroke as an exposure factor. However, there was no direct causal relationship between the subtypes of IS and FS. Our study provides theoretical support for shoulder care for patients with IS.

Efficacy of Acupuncture Treatment for Postprandial Distress Syndrome: A Systematic Review and Meta-Analysis.

Objective: This systematic review and meta-analysis was conducted to assess the efficacy of acupuncture treatment for postprandial distress syndrome (PDS). Methods: Search the Web of Science, the Cochrane Library, PubMed, and Embase databases with acupuncture randomized controlled trials for the treatment of patients with PDS. Strictly according to inclusion and exclusion quality assessment standards, the qualified ones are used to study the optimum extraction and data by two independent reviewers. Stata 15.0 software was used for meta-analysis. Result: We initially identified 63 studies, of which five (1253 participants) were eventually included in our analysis. There were 643 cases in the experimental group and 610 cases in the control group. Acupuncture had a significant effect on the total therapeutic effect (OTE) at week 4 (OR 4.74, 95% CI 02.88-7.83, Z = 6.10, P = 0 < 0.05). Significantly improved NDI (Nepean dyspepsia index) scores of PDS patients at week 4 (SMD 0.61, 95% CI 0.48 to 0.74). Significantly improved NDI scores in PDS patients at week 16 (SMD 0.49, 95% CI 0.27 to 0.71). After acupuncture treatment, the SID (dyspepsia symptom index) score of PDS patients decreased significantly at week 4 (SMD-0.52, 95% CI -0.73 to -0.32) and week 16 (SMD-0.59, 95% CI -0.81 to -0.36). Postprandial satiety scores (SMD-0.63, 95% CI -0.76 to -0.50) and early satiety scores (SMD-0.51, 95% CI -0.64 to -0.37) were also significantly lower at week 4 after acupuncture. Conclusion: This study highlighted that the acupuncture could significantly improve the overall therapeutic effect of PDS patients, alleviate the symptoms of postprandial fullness and early satiety, and improve the quality of life of patients. Our results supported that acupuncture was an effective therapeutic strategy for postprandial distress syndrome.

SAGE1: a Potential Target Antigen for Lung Cancer T-Cell Immunotherapy.

A fundamental understanding of cancer-specific antigens is crucial for successful T-cell immunotherapy. Sarcoma antigen 1 (SAGE1) is a cancer/testis antigen that has not yet been verified for T-cell immunotherapy applications. Here, we examined SAGE1 RNA expression and carried out IHC analyses, revealing that SAGE1 is expressed in 50% of non-small cell lung-cancer samples (n = 40). To verify the immunogenicity of SAGE1, we discovered a novel HLA-A*24:02 (HLA-A24)-restricted SAGE1 epitope (SAGE1597-606, VFSTAPPAFI) using mass spectrometry and identified SAGE1597-606-specific T-cell clones and T-cell receptors (TCR) from peripheral bloods of HLA-A24+ donors. The highest affinity TCR VF3 (KD = 4.3 µM) demonstrated the highest antitumor potency. Moreover, VF3-transduced T cells mediated the efficient killing of HLA-A24+/SAGE1+ tumor cells in vitro and effectively inhibited the growth of lung cancer xenografts in mice. Together, our data suggest that SAGE1 could be a target for T-cell immunotherapies against lung cancer, while its specific TCRs could be candidates for developing reagents to treat SAGE1+ tumors.

Identification of an HLA-A*24:02-restricted α-fetoprotein signal peptide-derived antigen and its specific T-cell receptor for T-cell immunotherapy.

Hepatocellular carcinoma (HCC) is the most common type of liver cancer with limited treatments. Asia has the highest HCC incidence rates; China accounts for over 50% of all HCC cases worldwide. T-cell receptor (TCR) -engineered T-cell immunotherapies specific for human leukocyte antigen (HLA) -A*02:01-restricted α-fetoprotein (AFP) peptide have shown encouraging results in clinics. HLA-A*24:02 is more common than HLA-A*02:01 in Asian countries, including China. Here we identified a novel HLA-A*24:02-restricted peptide KWVESIFLIF (AFP2-11 ) located in AFP signal peptide domain by mass spectrometric analysis of HLA-bound peptides from HepG2 cells. A TCR (KWV3.1) specific for AFP2-11 -HLA-A*24:02 was isolated from peripheral blood mononuclear cells of a healthy donor. The binding affinity of soluble KWV3.1 to its antigen was determined to be ~55 µm, within the affinity range of native TCRs for self-antigens. KWV3.1-transfected T cells could specifically activate and kill AFP2-11 pulsed T2-A24 cells and AFP+  HLA-A*24:02+ tumor cell lines, demonstrating that AFP2-11 can be naturally presented on the surface of AFP+ tumor cell lines. The newly identified antigenic peptide can provide a novel target for immunotherapeutic strategies for patients with AFP+  HLA-A*24:02+ HCC.

[Preparation of anti-human hemoglobin colloidal gold immunochromatography test strip].

Objective To develop the colloidal gold immunochromatography test strip for qualitatively detecting the hemoglobin (Hb) in human feces. Methods Mouse anti-human Hb monoclonal antibody SPR-5 marked by colloidal gold was coated in glass fiber membrane, and then the mouse anti-human Hb SP-5 monoclonal antibody and goat anti-mouse IgG were immobilized in testing (T) line and control (C) line located in nitrocellulose membranes, respectively. With this double antibody sandwich technique and immunochromatography test, the Hb antigen would react with both antibodies coated in the T line and C line and cause two colour reactions if the detected sample was positive, whereas the antigen-antibody combination and colour reaction only showed up in the C line when the sample was negative. Results The minimum detection limit of this test strip for human Hb was 21 ng/mL and no cross reactions were found in chick Hb, rabbit Hb, sheep Hb, pig Hb and cow Hb. Conclusion The test strips can improve the detection rate of fecal occult blood obviously and avoid false-positive results.

Engineering human MEK-1 for structural studies: A case study of combinatorial domain hunting.

Structural biology studies typically require large quantities of pure, soluble protein. Currently the most widely-used method for obtaining such protein involves the use of bioinformatics and experimental methods to design constructs of the target, which are cloned and expressed. Recently an alternative approach has emerged, which involves random fragmentation of the gene of interest and screening for well-expressing fragments. Here we describe the application of one such fragmentation method, combinatorial domain hunting (CDH), to a target which historically was difficult to express, human MEK-1. We show how CDH was used to identify a fragment which covers the kinase domain of MEK-1 and which expresses and crystallizes significantly better than designed expression constructs, and we report the crystal structure of this fragment which explains some of its superior properties. Gene fragmentation methods, such as CDH, thus hold great promise for tackling difficult-to-express target proteins.

Biochemical characterization of the active heterodimer form of human heparanase (Hpa1) protein expressed in insect cells.

The mammalian endoglycosidase heparanase (Hpa1) is primarily responsible for cleaving heparan sulphate proteoglycans (HSPGs) present on the basement membrane of cells and its potential for remodelling the extracellular matrix (ECM) could be important in embryonic development and tumour metastasis. Elevated expression of this enzyme has been implicated in various pathological processes including tumour cell proliferation, metastasis, inflammation and angiogenesis. The enzyme therefore represents a potential therapeutic target. Hpa1 protein is initially synthesized as an inactive 65 kDa proenzyme that is then believed to be subsequently activated by proteolytic cleavage to generate an active heterodimer of 8 and 50 kDa polypeptides. By analysis of a series of Hpa1 deletion proteins we confirm that the 8 kDa subunit is essential for enzyme activity. We present here for the first time an insect cell expression system used for the generation of large amounts of recombinant protein of high specific activity. Individual subunits were cloned into baculoviral secretory vectors and co-expressed in insect cells. Active secreted heterodimer protein was recovered from the medium and isolated by a one-step heparin-Sepharose chromatography procedure to give protein of >90% purity. The recombinant enzyme behaved similarly to the native protein with respect to the size of HS fragments liberated on digestion, substrate cleavage specificity and its preference for acidic pH. A significant amount of activity, however, was also detectable at physiological pH values, as measured both by an in vitro assay and by in vivo degradation of cell-bound heparan sulphate.