Fragment Antigen Research Articles

Probing the oligomeric re-assembling of bacterial fimbriae in vitro: a small-angle X-ray scattering and analytical ultracentrifugation study

Capsular antigen fragment 1 (Caf1) is an oligomeric protein consisting of 15 kDa monomeric subunits that are non-covalently linked through exceptionally strong and kinetically inert interactions into a linear polymer chain. It has been shown that after its thermal depolymerisation into unfolded monomeric subunits, Caf1 is able to efficiently repolymerise in vitro to reform its polymeric structure. However, little is known about the nature of the repolymerisation process. An improved understanding of this process will lead to the development of methods to better control the lengths of the repolymerised species, and ultimately, to better design of the properties of Caf1-based materials. Here we utilize small-angle X-ray scattering to estimate the size of Caf1 polymers during the first 24 h of the re-polymerisation process. Analytical ultracentrifugation measurements were also used to investigate the process post-24 h, where the rate of repolymerisation becomes considerably slower. Results show that in vitro polymerisation proceeds in a linear manner with no evidence observed for the formation of a lateral polymer network or uncontrolled aggregates. The rate of Caf1 in vitro repolymerisation was found to be concentration-dependent. Importantly, the rate of polymer growth was found to be relatively fast over the first few hours, before continuing at a dramatically slower rate. This observation is not consistent with the previously proposed step-growth mechanism of in vitro polymerisation of Caf1, where a linear increase in polymer length would be expected with time. We speculate how our observations may support the idea that the polymerisation process may be occurring at the ends of the chains with monomers adding sequentially. Our findings will contribute towards the development of new biomaterials for 3D cell culture and bio-printing.

Clinical Efficacy of Apatinib Combined with Gemcitabine on the Third Line or above in Patients with Advanced Lung Cancer and Its Effect on Serum Tumor Marker Levels

Objective . To analyze the clinical efficacy of apatinib combined with gemcitabine monotherapy for patients with third-line or higher advanced non-small cell lung cancer and its effect on serum tumor markers CA125, CEA, and CYERA21-1. Methods. A retrospective analysis was performed on 62 patients with stage III and IV third-line and above non-small cell lung cancer treated in the hospital from October 2016 to October 2018. Among them, 31 patients received gemcitabine monotherapy as a control group and 31 patients treated with apatinib combined with gemcitabine were used as the observation group, and the objective response rate (ORR), disease control rate (DCR), progression-free survival, overall survival of the two groups were compared and analyzed. Changes of serum tumor marker carbohydrate antigen (CA125), carcinoembryonic antigen (CEA), and cytokeratin fragment 19 (CYFRA21-1) in the two groups were observed before and after treatment. Results . After 8 weeks of treatment, the objective response rate of the observation group was 32.3%, the disease control rate was 80.6%, the objective response rate of the control group was 19.4%, the disease control rate was 58.1%, and the objective response rate and disease control rate of the observation group All were higher than the control group, and the difference was statistically significant ORR (χ 2 = 4.343, P 0.05). Conclusion. For patients with advanced non-small cell lung cancer, three-line or more treatment with apatinib combined with gemcitabine compared with gemcitabine single-drug regimen has improved short-term and long-term efficacy, and most of the adverse reactions are tolerable and safe.

Research progress on the molecular mechanisms of hepatic metastasis in lung cancer: a narrative review.

The liver is one of the most common sites of metastatic spread of lung cancer, and the process of metastasis is regulated by many factors. A number of genes, including multiple tumor suppressor 1 (mts1), p120 catenin, and CT45A1, increase the possibility of hepatic metastasis in lung cancer, whereas Tip30/CC3, CUL5, and SOCS3 expression in lung tumors inhibit tumor metastasis. microRNAs (miRNAs), such as miRNA-126, miRNA-338, and miRNA-218, can affect the metastasis of lung cancer cells to the liver. The D114-Notch signaling pathway can inhibit liver metastasis in small cell lung cancer. Serum tumor markers cytokeratin 19 fragment antigen 21-1 and neuron-specific enolase (NSE) are closely related to the risk of hepatic metastasis in lung cancer. Based on previously published literature, we found that the metastasis and invasion of lung cancer to the liver are determined by molecular factors. Therefore, the selective identification and intervention of these erroneous signals can predict early lung cancer metastasis to the liver. In this review article, we describe the mechanisms and influencing factors (genes, signal pathways, chemicals, proteins, miRNAs) of hepatic metastasis in lung cancer. We hope to provide a summary of the evidence for the mechanisms by which related genes or proteins affect the malignancy of liver metastasis from lung cancer so that doctors and researchers can improve treatment options.

Lateral flow biosensor for universal detection of various targets based on hybridization chain reaction amplification strategy with pregnancy test strip

Abstract Pregnancy test strip is currently one of the most mature and widely used biomedical test products. In this study, based on its simplicity and rapidity, the detecting target-human chorionic gonadotropin (hCG) is coupled with different hybridization probes, while the aptamer is linked with the initiator used for hybridization chain reaction (HCR), which can construct a signal amplification platform based on HCR. The method converts the detection of other biochemical molecules into the detection of hCG signal, and combines the magnetic separation technology to realize the detection of various targets with pregnancy test strip. The rapid quantitative detection can be achieved with smartphone App and portable detection device. Under the optimal experiment conditions, the method can realize rapid and sensitive detection of adenosine triphosphate, thrombin, carcinoembryonic antigen and DNA fragment in human serum, and the visual detection limits can reach 100 pM, 0.2 pM, 5 ng/mL and 5 pM, respectively. The design has low cost, simple operation, and good sensitivity and selectivity for target detection. Based on this design, the detection of multiple analytes can be implemented with different aptamer sequences, thereby broadening the application of commercial pregnancy test strip in the biomedical testing field.

Novel electrochemical biosensor based on Exo III-assisted digestion of dsDNA polymer from hybridization chain reaction in homogeneous solution for CYFRA 21-1 DNA assay

A novel electrochemical biosensing strategy was proposed to detect cytokeratin fragment antigen 21-1 (CYFRA 21-1) DNA based on Exo III-assisted digestion of dsDNA polymer (EADDP) from hybridization chain reaction (HCR). Primarily, the presence of target can drive a catalytic hairpin assembly (CHA) reaction, which was aimed to achieve target recognition and circulation. Then the HCR can be triggered for further signal amplification and generate long dsDNA polymer with signal tags. Subsequently, the introduction of Exo III can digest the long dsDNA polymer to produce large amounts of double signal fragments (DSFs). The above experiments were all carried out in homogeneous solution. Finally, the released DSF can be captured onto the electrode directly by capture probe (CP) and a highly amplified electrochemical signal can be detected. The EADDP in homogeneous solution circumvented complex solid-liquid interface reaction and tedious operation steps on electrode. Besides, one target can be converted into abundant DSFs, which greatly improved the sensitivity. This biosensor exhibited a low detection limit (0.0348 fM) and wide linear range (5 fM ∼ 50 nM) for CYFRA 21-1 DNA biosensing with reliable specificity and stability.

Shedding light on TCR activation

Immune Signaling In mammals, T cells recognize fragments of pathogen and other alien peptide antigens using T cell receptors (TCRs). Loaded TCRs then interact with the polymorphic cell surface proteins of the major histocompatibility complexes of antigen-presenting cells, activating killing processes. O'Donoghue et al. engineered CD4+ T cells with an optogenetic chimeric antigen receptor stimulated by blue light. This technique provides an experimental on–off switch with which to precisely control the TCR-signaling machinery. The authors could distinguish between the signals induced by self and foreign antigens, the latter inducing CD69, a marker of T cell activation. Proc. Natl. Acad. Sci. U.S.A. 118 , e2019285118 (2021).

Electrochemiluminescence immunosensor for cytokeratin fragment antigen 21-1 detection using electrochemically mediated atom transfer radical polymerization

The cytokeratin fragment antigen 21-1 (CYFRA 21-1) protein is a critical tumor biomarker tightly related to non-small cell lung cancer (NSCLC). Herein, we prepared an effective electrochemiluminescence (ECL) immunosensor for CYFRA 21-1 detection using electrochemically mediated atom transfer radical polymerization (eATRP). The CYFRA 21-1 antigen was fixed on the electrode surface by constructing a sandwich type antibody-antigen-antibody immune system. The sensitivity of ECL was improved by using the eATRP reaction. In this method, eATRP was applied to CYFRA 21-1 detection antibody with N-acryloyloxysuccinimide as functional monomer. This is the first time that ECL and eATRP signal amplification technology had been combined. Under the optimized testing conditions, the immunosensor showed a good linear relation in the range from 1 fg mL−1 to 1 μg mL−1 at a limit of detection of 0.8 fg mL−1 (equivalent to ~ 134 molecules in a 10 μL sample). The ECL immunosensing system based on eATRP signal amplification technology provided a new way for rapid diagnosis of lung cancer by detecting CYFRA 21-1.Graphical abstractThe paper prepared an electrochemiluminescence biosensor for ultrasensitive detection of CYFRA 21-1 via eATRP signal amplification strategy, which had the advantages of high sensitivity, reproducibility, and held potential prospect for analysis of low-abundance.

Comparison of serum tumor markers combined with dual-source CT in the diagnosis of lung cancer.

The purpose of this study is to explore the clinical value of serum tumor markers combined with dual-source CT scanning in the diagnosis of lung cancer. One hundred-two patients with lung cancer (malignant tumor group), 50 patients with benign lesions (benign control group) and 50 healthy patients (normal control group) were selected as the research objects. The levels of serum carcinoembryonic antigen (CEA), cytokeratin 19 fragment antigen 21-1 (CYFRA21-1) and gastrin releasing peptide precursor 31-98 (Pro-GRP31-98) were detected using the electrochemiluminescence and enzyme-linked immunoassay in three groups of people. Simultaneously, Siemens' second-generation dual-source CT (Siemens AG, Munich, Germany) is used to scan the lungs of patients with lung cancer and benign lesions. Retrospective statistical analysis is utilized to explore the clinical value of serum tumor markers combined with dual-source CT examination in the diagnosis of lung cancer. The levels of serum CEA, CYFRA21-1 and Pro-GRP31-98 in lung cancer patients were significantly higher than those in the benign control group and normal control group, and the difference was statistically significant (P<0.01). There was no statistical difference between the benign control group and the normal control group (P>0.05). The levels of serum CEA, CYFRA21-1, Pro-GRP31-98 had no significant correlation with the age, sex, and tumor site of lung cancer patients (P>0.05). However, the levels of serum CEA, CYFRA21-1, Pro-GRP31-98 have an obvious correlation with tumor size, clinical stage, histological type, combined pleural effusion, recurrence and metastasis after treatment (P<0.05). CT scans of 102 lung cancer patients showed 35 cases of central type, 67 cases of peripheral type, 71 cases with tumor diameter >5cm, 31 cases of ≤5cm and 67 cases of lymph node metastasis. The sensitivities of CEA, CYFRA21-1, Pro-GRP31-98, and dual-source CT in the diagnosis of lung cancer were 64.70%, 60.11%, 56.86% and 77.45%, and the accuracy was 75.00%, 72.37%, 69.74% and 82.89%, respectively. Compared with a single examination, the sensitivity and accuracy of combined examination for the diagnosis of lung cancer were significantly improved, which were 95.10% and 92.76%, respectively. Joint examinations can effectively improve the rate of lung cancer diagnosis.

Anti-SARS-CoV-2 Antibodies Within IVIg Preparations: Cross-Reactivities With Seasonal Coronaviruses, Natural Autoimmunity, and Therapeutic Implications

Introduction: Cross-reactivity to SARS-CoV-2 antigenic peptides has been detected on T-cells from pre-pandemic donors due to recognition of conserved protein fragments within members of the coronavirus's family. Further, preexisting antibodies recognizing SARS-CoV-2 with conserved epitopes in the spike region have been now seen in uninfected individuals. High-dose Intravenous Immunoglobulin (IVIg), derived from thousands of healthy donors, contains natural IgG antibodies against various antigens which can be detected both within the IVIg preparations and in the serum of IVIg-receiving patients. Whether IVIg preparations from pre-pandemic donors also contain antibodies against pre-pandemic coronaviruses or autoreactive antibodies that cross-react with SARS-CoV-2 antigenic epitopes, is unknown.Methods: 13 samples from 5 commercial IVIg preparations from pre-pandemic donors (HyQvia (Baxalta Innovations GmbH); Privigen (CSL Behring); Intratect (Biotest AG); IgVena (Kedrion S.p.A); and Flebogamma (Grifols S.A.) were blindly screened using a semi-quantitative FDA-approved and validated enzyme-linked immunosorbent assay (ELISA) (Euroimmun, Lubeck, Germany).Results: Nine of thirteen preparations (69.2%), all from two different manufactures, were antibody-positive based on the defined cut-off positivity (index of sample OD to calibrator OD > 1.1). From one manufacturer, 7/7 lots (100%) and from another 2/3 lots (67%), tested positive for cross-reacting antibodies. 7/9 of the positive preparations (77%) had titers as seen in asymptomatically infected individuals or recent COVID19-recovered patients, while 2/9 (23%) had higher titers, comparable to those seen in patients with active symptomatic COVID-19 infection (index > 2.2).Conclusion: Pre-pandemic IVIg donors have either natural autoantibodies or pre-pandemic cross-reactive antibodies against antigenic protein fragments conserved among the “common cold” - related coronaviruses. The findings are important in: (a) assessing true anti-SARS-CoV-2-IgG seroprevalence avoiding false positivity in IVIg-receiving patients; (b) exploring potential protective benefits in patients with immune-mediated conditions and immunodeficiencies receiving acute or chronic maintenance IVIg therapy, and (c) validating data from a recent controlled study that showed significantly lower in-hospital mortality in the IVIg- treated group.

Electrochemical ultrasensitive detection of CYFRA21-1 using Ti3C2Tx-MXene as enhancer and covalent organic frameworks as labels.

The concentration level of cytokeratin fragment antigen 21-1 (CYFRA21-1) can be used as an important indicator for predicting non-small cell lung cancer (NSCLC). Here, a sandwich-type electrochemical immunosensor for ultrasensitive detection of CYFRA21-1 is developed. The sensor based on a combination of gold nanoparticle (AuNPs) decorated Ti3C2Tx-MXene (Au-Ti3C2Tx) as the substrate enhancer, and toluidine blue (TB) modified AuNPs doped covalent organic framework (COF) polymer as the signal tag (TB-Au-COF). The Au-Ti3C2Tx is used to capture numerous primary antibodies and accelerate the electron transfer rate of the substrate, while the TB-Au-COF can be applied to provide a large number of signal units TB and secondary antibodies. These features of composites endow the proposed immunosensor with high sensitivity and current response to CYFRA21-1. Under optimum conditions, the immunosensor offers a wide current response for CYFRA21-1 from 0.5-1.0 × 104pg·mL-1 with a detection limit of 0.1pg·mL-1. Furthermore, the biosensing platform can be applied for CYFRA21-1 detection to analyze real serum samples, providing an effective and useful avenue for the applicability of Au-Ti3C2Tx and TB-Au-COF composite materials in biosensing field.

Seroprevalence of Toxoplasma gondii assayed using Rapid Diagnostic Tests among Residents in Three Counties Adjacent to The Demilitarized Zone, Korea

Toxoplasma gondii seroprevalence have been rapidly increasing in some parts of Korea. We analyzed prevalence of anti-Toxoplasma gondii antibodies, using a rapid diagnostic test (RDT), in the sera of 552 residents in Ganghwa-gun, 661 ones in Cheorwon-gun, and 305 ones in Goseong-gun, Korea in 2019. IgG/IgM RDT mounted with recombinant fragment of major surface antigen (SAG1), glutathione-S-transferase-linker-SAG1A, were applied to the sera. IgG seroprevalence was 28.1% in Ganghwa-gun, 19.5% in Cheorwon-gun and 35.7% in Goseong-gun. Odds ratios comparing Cheorwon vs Ganghwa was 0.63 (P=0.001) and Goesong versus Ganghwa was 1.47 (P=0.01) adjusting age and sex. Goseong had highest seroprevalence among the 3 counties both in crude rates and logistic regression. Although Cheorwon and Goseong are adjacent to the demilitarized zone (DMZ) in Korea, seroprevalence rate was much higher in Goseong. Further investigation on other DMZ-closed areas is necessary whether they have high prevalence rates compared to the other areas. T. gondii prevalence in Korea is still persists; proper health policy should be established.

Modular vaccine platform based on the norovirus-like particle

BackgroundVirus-like particle (VLP) vaccines have recently emerged as a safe and effective alternative to conventional vaccine technologies. The strong immunogenic effects of VLPs can be harnessed for making vaccines against any pathogen by decorating VLPs with antigens from the pathogen. Producing the antigenic pathogen fragments and the VLP platform separately makes vaccine development rapid and convenient. Here we decorated the norovirus-like particle with two conserved influenza antigens and tested for the immunogenicity of the vaccine candidates in BALB/c mice.ResultsSpyTagged noro-VLP was expressed with high efficiency in insect cells and purified using industrially scalable methods. Like the native noro-VLP, SpyTagged noro-VLP is stable for months when refrigerated in a physiological buffer. The conserved influenza antigens were produced separately as SpyCatcher fusions in E. coli before covalent conjugation on the surface of noro-VLP. The noro-VLP had a high adjuvant effect, inducing high titers of antibody production against the antigens presented on its surface.ConclusionsThe modular noro-VLP vaccine platform presented here offers a rapid, convenient and safe method to present various soluble protein antigens to the immune system for vaccination and antibody production purposes.

An In-silico Approach and Experimental Analysis Combination: Two Strategies for Selecting the third Extracellular Domain (D-EC3) of Human CD133 Marker as a Target for Detection of Cancer Stem Cells.

The selection of the appropriate fragment of the cell surface receptors as an antigen is significant for the production of antibodies. CD133, as a suitable biomarker candidate in the cancer stem cells (CSCs), is a glycosylated protein. The antibodies used for analyzing it recognize glycosylated epitopes of CD133. Since the glycosylated motifs have a dynamic nature over the lifetime of a protein, they limit the detection of CD133. In this study, to access a specific antibody against the antigenic, accessible, and non-glycosylated fragment of the native CD133, we performed an in-silico analysis. Then, we expressed the third domain (D-EC3) (serine641-leucine710) in E. coli BL21 (DE3), then the purified recombinant antigen immunized BALB/c mice. Finally, the dignity of an epitope of pure recombinant antigen has been approved by the interactions of antibody and antigen with the use of mice immunized sera via ELISA and flow cytometry experimentation. The results showed that the selected non-glycosylated fragment can compete well with the commercial antibody against the glycosylated epitopes to identify the native cell surface markers. The results can be considered for diagnosis and target therapy development of CD133+ cancer cells.

Hydrogels of engineered bacterial fimbriae can finely tune 2D human cell culture.

Demand continues to grow for biomimetic materials able to create well-defined environments for modulating the behaviour of living cells in culture. Here, we describe hydrogels based upon the polymeric bacterial fimbriae protein capsular antigen fragment 1 (Caf1) that presents tunable biological properties for enhanced tissue cell culture applications. We demonstrate how Caf1 hydrogels can regulate cellular functions such as spreading, proliferation and matrix deposition of human dermal fibroblast cells (hDFBs). Caf1 hydrogels exploring a range of mechanical properties were prepared using copolymers featuring controlled compositions of inert wild-type Caf1 subunits and a mutant subunit displaying the RGDS peptide motif. The hydrogels showed excellent cytocompatibility with hDFBs and the ability to modulate both cell morphology and matrix deposition. Interestingly, Caf1 hydrogels displaying faster stress relaxation were demonstrated to show the highest metabolic activities of growing cells in comparison with other Caf1 hydrogel formulations. The stiffest Caf1 hydrogel impacted cellular morphology, inducing alignment of the cells. This work is significant as it clearly indicates that Caf1-based hydrogels offer tuneable biochemical and mechanical substrates conditions suitable for cell culture applications.

A novel electrochemical lung cancer biomarker cytokeratin 19 fragment antigen 21-1 immunosensor based on Si3N4/MoS2 incorporated MWCNTs and core-shell type magnetic nanoparticles.

Lung cancer isone ofdeadliest and most life threateningcancer types. Cytokeratin 19 fragment antigen 21-1 (CYFRA 21-1) is a significant biomarker for the diagnosis of non-small cell lung cancer (NSCLC). Due to these reasons, a novel electrochemical immunosensor based on a silicon nitride (Si3N4)-molybdenum disulfide (MoS2) composite on multi-walled carbon nanotubes (Si3N4/MoS2-MWCNTs) as an electrochemical sensor platform and core-shell type magnetic mesoporous silica nanoparticles@gold nanoparticles (MMSNs@AuNPs) as a signal amplifier was presented for CYFRA21-1 detection in this study. Capture antibody (Ab1) immobilization on a Si3N4/MoS2-MWCNT modified glassy carbon electrode (Si3N4/MoS2-MWCNTs/GCE) was firstly successfully performed by stable electrostatic/ionic interactions between the -NH2 groups of the capture antibody and the polar groups of Si3N4/MoS2. Then, specific antibody-antigen interactions between the electrochemical sensor platform and the signal amplifier formed a novel voltammetric CYFRA21-1 immunosensor. The prepared composite materials and electrochemical sensor surfaces were characterized by transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). A linearity range of 0.01-1.0 pg mL-1 and a low detection limit (LOD) of 2.00 fg mL-1 were also obtained for analytical applications. Thus, the proposed immunosensor based on Si3N4/MoS2-MWCNTs and MMSNs@AuNPs has great potential for medical diagnosis of lung cancer.

Ultrasensitive electrochemical immunosensor via RAFT polymerization signal amplification for the detection of lung cancer biomarker

Abstract Cytokeratin fragment antigen 21–1 (CYFRA21-1), a characteristic biomarker of non-small cell lung cancer (NSCLC), its abundance is closely related to the diagnosis and prognosis of the disease. Thus, an ultrasensitive electrochemical immunosensor for the detection of CYFRA21-1 was designed by exploiting reversible addition-fragmentation chain transfer (RAFT) polymerization signal amplification strategy. Briefly, graphene oxide (GO) was immobilized on a glassy carbon electrode (GCE) surface to bind a CYFRA21-1 primary antibody (Ab1) via an amide bond, allowing the analyte CYFRA21-1 to be selectively anchored to Ab1 by specific antigen-antibody interactions. Then, the chain transfer agent-conjugated a CYFRA21-1 secondary antibody was attached to the CYFRA21-1, forming a sandwich-like immunocomplex that can be linked to numerous monomers through RAFT polymerization. Accordingly, a high-density electroactive polymer chain was then grafted onto the immunocomplex, dramatically amplifying the electrochemical signal generated by the immobilized CYFRA21-1. Using the strategy, the current response increased linearly with the logarithm of the CYFRA21-1 concentration in the range of 0.5 fg mL−1 to 10 pg mL−1 (R2 = 0.998), with a low detection limit of 0.14 fg mL−1 (S/N = 3). Furthermore, the CYFRA21-1 concentrations in clinical serum samples analyzed by our method were consistent with those obtained by magnetic particle chemiluminescence assay (MPCA) method. Thus, owing to its high sensitivity, environmental benignity, and low cost, this strategy shows great potential for clinical diagnosis and analysis.

The Immunogenic and Immunoprotective Activities of Recombinant Chimeric T. gondii Proteins Containing AMA1 Antigen Fragments.

Toxoplasmosis, one of the most common parasitoses worldwide, is potentially dangerous for individuals with a weakened immune system, but specific immunoprophylaxis intended for humans is still lacking. Thus, efforts have been made to create an efficient universal vaccine for both animals and humans to overcome the shortcomings of currently used treatment methods and protect all hosts against toxoplasmosis. The current work represents a relatively new approach to vaccine development based on recombinant chimeric Toxoplasma gondii antigens. In the present research, three tetravalent chimeric proteins containing different portions of the parasite’s AMA1 antigen—AMA1domain I-SAG2-GRA1-ROP1L (ANSGR), AMA1domains II, III-SAG2-GRA1-ROP1L (ACSGR) and AMA1full protein-SAG2-GRA1-ROP1L (AFSGR)—were tested for their immunogenic and immunoprotective capacities. All tested proteins were immunogenic, as evidenced by the triggering of specific humoral and cellular immune responses in vaccinated C3H/HeOuJ mice, defined by the production of specific IgG (IgG1/IgG2a) antibodies in vivo and synthesis of key Th1/Th2 cytokines by Toxoplasma lysate antigen-stimulated splenocytes in vitro. Although all tested preparations provided partial protection against chronic toxoplasmosis in immunized and T. gondii-challenged mice, the intensity of the generated immunoprotection depended on the fragment of the AMA1 antigen incorporated into the chimeric antigen’s structure.

Mechanism of action of Panax notoginoside against lung cancer in mice based on response to CTSB gene

BackgroundThis study aimed to investigate the mechanism of action of Panax notoginoside (PNS) against lung cancer and inhibition of lung cancer cell proliferation by the drug at different concentrations in a mouse model, considering the cathepsin B (CTSB) gene as a target.MethodsThe mice were randomly assigned into the following five groups: normal control, tumor-bearing, low-dose Panax notoginoside (TSPN), medium-dose TSPN, and high-dose TSPN. All mice were treated with physiological saline or TSPN at different concentrations for 28 days consecutively by gavage. The tumor size was measured, the tumor growth was observed, and the survival curve was drawn. At different time points, the expression of the CTSB gene was detected using quantitative fluorescent polymerase chain reaction, Western blot analysis, and indirect immunofluorescence. The serum indices, such as carcinoembryonic antigen (CEA), neuron-specific enolase (NSE), and Soluble fragment of cytokeratin 19 (CYFRA21), were detected by enzyme-linked immunosorbent assay.ResultsIn vivo, PNS could directly inhibit the expression of the CTSB gene in tumors of mice, limit tumor growth, and alter tumor-related indices, such as CEA, NSE, and CYFRA21 levels, in the serum to different extents simultaneously.ConclusionCTSB gene was closely related to the pathogenesis of lung cancer. PNS could act on the CTSB gene, downregulate the expression of CTSB in lung cancer cells, inhibit the proliferation and invasion of tumors, and prolong the survival period.

Cytotoxic activity of human dendritic cells induces RIPK1-dependent cell death

Dendritic cells (DCs), as potent phagocytes engulf dead cells and present peptide fragments of tumor antigens or pathogens derived from infected cells to naïve CD8+ T-lymphocytes. Dendritic cells can also induce apoptosis in target cells, thus getting an opportunity to sample their microenvironment. Here, we present that the supernatants of LPS- or CL075-activated DCs induced cell death in different cell lines, but during the differentiation to mature DCs, they lost their cytotoxic potential. Dexamethasone-pre-treated tolerogenic DCs induced less intensive death indicating that the tissue microenvironment can downregulate DC-mediated killing. Exploring the signaling of DC-induced cell death, we observed that the supernatant of activated DCs induced TNF-dependent cell death, since TNF antagonist blocked the cytotoxic activity of DCs, contrary to inhibitors of Fas and TRAIL receptors. We identified that the DC-induced killing is at least partially a RIPK1-dependent process, as RIPK1 positive target cells were more susceptible to DC-induced cell death than their RIPK1 deficient counterparts. Moreover, both the elevated phosphorylation of RIPK1 and the increase in RIPK1-caspase-8 interaction in target cells suggest that RIPK1-mediated signals contribute to DC supernatant-induced cell death. We also proved that the cytotoxic activity of DC-derived supernatant induced apoptosis in the target cells and not necroptosis, as it was completely abrogated with the pan caspase inhibitor (Z-VAD), while the necroptosis inhibitor (Nec-1) had no effect.Our work revealed that the supernatant of activated DCs induces the apoptosis of target cells in a RIPK1-dependent manner. This phenomenon could be relevant for the initiation of cross-presentation and may broaden the plethora of cytotoxic mechanisms acting against tumor cells.

Peripheral Blood Leukocyte N6-methyladenosine is a Noninvasive Biomarker for Non-small-cell Lung Carcinoma.

BackgroundN6-methyladenosine (m6A) triggers a new layer of epi-transcription. However, the potential noninvasive screening and diagnostic value of peripheral blood m6A for cancer are still unknown. Here, we intend to investigate whether leukocyte m6A can be a novel biomarker for non-small-cell lung cancer (NSCLC).Materials and MethodsPeripheral blood was collected from 119 NSCLC patients and 74 age-matched healthy controls. Total RNA was isolated from leukocytes for m6A measurement, and clinical information of participants was reviewed. The sensitivity, specificity, and area under the curve (AUC) of m6A for cancer diagnosis were evaluated by the receiver-operating characteristic (ROC) curve analysis. Flow cytometry and the Human Protein Atlas (HPA) database were used to characterize m6A in leukocyte differentials. Pearson’s correlation was applied to indicate the relationship between m6A level and hematology variables. qPCR and bioinformatic analysis were used to identity the expression of m6A regulators in leukocyte.ResultsLeukocyte m6A was significantly elevated in 119 NSCLC patients compared with 74 healthy controls (P<0.001). We did not find significant association between m6A and age or gender. Elevated m6A level in NSCLC was associated with tumor stage (P<0.05) and tumor differentiation (P<0.05), and was significantly reduced after surgery (P<0.01). ROC curve analysis revealed that leukocyte m6A could significantly discriminate patients with lung adenocarcinoma (LUAD) (AUC=0.736, P<0.001) and lung squamous cell carcinoma (LUSC) (AUC=0.963, P<0.001) from healthy individuals. m6A displayed superior sensitivity (100%) and specificity (85.7%) for LUSC than squamous cell carcinoma (SCC) antigen and cytokeratin fragment 211 (Cyfra211). Flow cytometry analysis showed m6A modification was mainly localized on T cells and monocytes among leukocyte differentials. Leukocyte m6A was positively correlated with the number of lymphocytes and negatively correlated with monocytes in NSCLC but not in healthy controls. qPCR and bioinformatic analysis showed that elevated leukocyte m6A in NSCLC was caused by upregulated methyltransferase complex and downregulated FTO and ALKBH5.ConclusionLeukocyte m6A represents a potential noninvasive biomarker for NSCLC screening, monitoring and diagnosis.