Identification of an HLA-A*11:01-restricted neoepitope of mutant PIK3CA and its specific T cell receptors for cancer immunotherapy targeting hotspot driver mutations.

Hotspot driver mutations presented by human leukocyte antigens might be recognized by anti-tumor T cells. Based on their advantages of tumor-specificity and immunogenicity, neoantigens derived from hotspot mutations, such as PIK3CAH1047L, may serve as emerging targets for cancer immunotherapies. NetMHCpan V4.1 was utilized for predicting neoepitopes of PIK3CA hotspot mutation. Using in vitro stimulation, antigen-specific T cells targeting the HLA-A*11:01-restricted PIK3CA mutation were isolated from healthy donor-derived peripheral blood mononuclear cells. T cell receptors (TCRs) were cloned using single-cell PCR and sequencing. Their functionality was assessed through T cell activation markers, cytokine production and cytotoxic response to cancer cell lines pulsed with peptides or transduced genes of mutant PIK3CA. Immunogenic mutant antigens from PIK3CA and their corresponding CD8+ T cells were identified. These PIK3CA mutation-specific CD8+ T cells were subsequently enriched, and their TCRs were isolated. The TCR clones exhibited mutation-specific and HLA-restricted reactivity, demonstrating varying degrees of functional avidity. Identified TCR genes were transferred into CD8+ Jurkat cells and primary T cells deficient of endogenous TCRs. TCR-expressing cells demonstrated specific recognition and reactivity against the PIK3CAH1047L peptide presented by HLA-A*11:01-expressing K562 cells. Furthermore, mutation-specific TCR-T cells demonstrated an elevation in cytokine production and profound cytotoxic effects against HLA-A*11:01+ malignant cell lines harboring PIK3CAH1047L. Our data demonstrate the immunogenicity of an HLA-A*11:01-restricted PIK3CA hotspot mutation and its targeting therapeutic potential, together with promising candidates of TCR-T cell therapy.

Neutrophil depletion attenuates antibody-mediated rejection in a renal transplantation mouse model.

Antibody-mediated rejection (AMR) can cause graft failure following renal transplantation. Neutrophils play a key role in AMR progression, but the exact mechanism remains unclear. We investigated the effect of neutrophils on AMR in a mouse kidney transplantation model. The mice were divided into five groups: syngeneic transplantation (Syn), allograft transplantation (Allo), and three differently treated AMR groups. The AMR mouse model was established using skin grafts to pre-sensitize recipient mice. Based on the AMR model, Ly6G-specific monoclonal antibodies were administered to deplete neutrophils (NEUT-/- + AMR) and TACI-Fc was used to block B-cell-activating factor (BAFF)/a proliferation-inducing ligand (APRIL) signaling (TACI-Fc + AMR). Pathological changes were assessed using hematoxylin-eosin and immunohistochemical staining. Banff values were evaluated using the Banff 2015 criteria. Donor-specific antibody (DSA) levels were assessed using flow cytometry, and BAFF and APRIL concentrations were measured using ELISA. Compared to the Syn and Allo groups, a significantly increased number of neutrophils and increased C4d and IgG deposition were observed in AMR mice, accompanied by elevated DSA levels. Neutrophil depletion inhibited inflammatory cell infiltration and reduced C4d and IgG deposition. Neutrophil depletion significantly decreased DSA levels after transplantation and suppressed BAFF and APRIL concentrations, suggesting a mechanism for attenuating AMR-induced graft damage. Similar results were obtained after blockading BAFF/APRIL using a TACI-Fc fusion protein. In summary, neutrophil infiltration increased in the AMR mouse renal transplantation model. Neutrophil depletion or blockading the BAFF/APRIL signaling pathway significantly alleviated AMR and may provide better options for the clinical treatment of AMR.

A non-RBM targeted RBD specific antibody neutralizes SARS-CoV-2 inducing S1 shedding.

Potent neutralizing antibodies (Abs) have been proven with therapeutic efficacy for the intervention against SARS-CoV-2. Majority of these Abs function by directly interfering with the virus entry to host cells. Here, we identified a receptor binding domain (RBD) specific monoclonal Ab (mAb) 82A6 with efficient neutralizing potency against authentic SARS-CoV-2 virus. As most Abs targeting the non-receptor binding motif (RBM) region, 82A6 was incapable to block the RBD-ACE2 interaction. In particular, it actively promoted the S1 subunit shedding from the S protein, which may lead to effective reduction of intact SARS-CoV-2 viruses. Importantly, it could block potential syncytia formation associated with post-infectious cell surface expression of S proteins. Our study evidenced a RBD specific Ab with unique beneficial efficacy against SARS-CoV-2 infection, which might bring informative significance to understand the collective effects of neutralizing Abs elicited in COVID-19 patients.

Fully human monoclonal antibodies to TRAIL-R1 enhance TRAIL-induced apoptosis via activation of caspase-8 pathway.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) or agonistic antibodies targeting TRAIL-receptors (TRAIL-Rs) can selectively induce apoptosis in cancer cells. However, they have limited antitumor efficacy in clinical trials. We previously generated ten fully human monoclonal Abs to TRAIL-receptor type 1 (TR1-mAbs) using immunospot array assay on a chip (ISAAC technology). We found that the TR1-mAbs exhibited different effects on TRAIL-induced apoptosis (enhanced or blocked apoptosis). Here, we further demonstrated that some mAbs competed with TRAIL for binding to TRAIL-R1 expressed on tumor cells that blocked TRAIL-induced apoptosis (B-TR1-Ab), whereas others did not compete with TRAIL that enhanced TRAIL-induced apoptosis (E-TR1-Ab). Combination of E-TR1-Ab (TR1-419) with TRAIL leads to enhanced antitumor activity in various tumor cells in vitro. E-TR1-419 and TRAIL could cooperate to upregulate the mRNA expression and protein levels of TRAIL-R1 and to promote caspase-8 cleavage and increased JNK phosphorylation. Our results suggest that combining E-TR1 Ab with TRAIL could provide a new therapeutic strategy for tumor immunotherapies.

A chimeric antigen receptor for TRAIL-receptor 1 induces apoptosis in various types of tumor cells.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and its associated receptors (TRAIL-R/TR) are attractive targets for cancer therapy because TRAIL induces apoptosis in tumor cells through TR while having little cytotoxicity on normal cells. Therefore, many agonistic monoclonal antibodies (mAbs) specific for TR have been produced, and these induce apoptosis in multiple tumor cell types. However, some TR-expressing tumor cells are resistant to TR-specific mAb-induced apoptosis. In this study, we constructed a chimeric antigen receptor (CAR) of a TRAIL-receptor 1 (TR1)-specific single chain variable fragment (scFv) antibody (TR1-scFv-CAR) and expressed it on a Jurkat T cell line, the KHYG-1 NK cell line, and human peripheral blood lymphocytes (PBLs). We found that the TR1-scFv-CAR-expressing Jurkat cells killed target cells via TR1-mediated apoptosis, whereas TR1-scFv-CAR-expressing KHYG-1 cells and PBLs killed target cells not only via TR1-mediated apoptosis but also via CAR signal-induced cytolysis, resulting in cytotoxicity on a broader range if target cells than with TR1-scFv-CAR-expressing Jurkat cells. The results suggest that TR1-scFv-CAR could be a new candidate for cancer gene therapy.

Dynamic Survival Risk Prognostic Model and Genomic Landscape for Atypical Teratoid/Rhabdoid Tumors: A Population-Based, Real-World Study.

BACKGROUND: An atypical teratoid/rhabdoid tumor (AT/RT) is an uncommon and aggressive pediatric central nervous system neoplasm. However, a universal clinical consensus or reliable prognostic evaluation system for this malignancy is lacking. Our study aimed to develop a risk model based on comprehensive clinical data to assist in clinical decision-making. METHODS: We conducted a retrospective study by examining data from the Surveillance, Epidemiology, and End Results (SEER) repository, spanning 2000 to 2019. The external validation cohort was sourced from the Children's Hospital Affiliated to Chongqing Medical University, China. To discern independent factors affecting overall survival (OS) and cancer-specific survival (CSS), we applied Least Absolute Shrinkage and Selection Operator (LASSO) and Random Forest (RF) regression analyses. Based on these factors, we structured nomogram survival predictions and initiated a dynamic online risk-evaluation system. To contrast survival outcomes among diverse treatments, we used propensity score matching (PSM) methodology. Molecular data with the most common mutations in AT/RT were extracted from the Catalogue of Somatic Mutations in Cancer (COSMIC) database. RESULTS: The annual incidence of AT/RT showed an increasing trend (APC, 2.86%; 95% CI:0.75-5.01). Our prognostic study included 316 SEER database participants and 27 external validation patients. The entire group had a median OS of 18 months (range 11.5 to 24 months) and median CSS of 21 months (range 11.7 to 29.2). Evaluations involving C-statistics, DCA, and ROC analysis underscored the distinctive capabilities of our prediction model. An analysis via PSM highlighted that individuals undergoing triple therapy (integrating surgery, radiotherapy, and chemotherapy) had discernibly enhanced OS and CSS. The most common mutations of AT/RT identified in the COSMIC database were SMARCB1, BRAF, SMARCA4, NF2, and NRAS. CONCLUSIONS: In this study, we devised a predictive model that effectively gauges the prognosis of AT/RT and briefly analyzed its genomic features, which might offer a valuable tool to address existing clinical challenges.

SMAD4 depletion contributes to endocrine resistance by integrating ER and ERBB signaling in HR + HER2- breast cancer.

Endocrine resistance poses a significant clinical challenge for patients with hormone receptor-positive and human epithelial growth factor receptor 2-negative (HR + HER2-) breast cancer. Dysregulation of estrogen receptor (ER) and ERBB signaling pathways is implicated in resistance development; however, the integration of these pathways remains unclear. While SMAD4 is known to play diverse roles in tumorigenesis, its involvement in endocrine resistance is poorly understood. Here, we investigate the role of SMAD4 in acquired endocrine resistance in HR + HER2- breast cancer. Genome-wide CRISPR screening identifies SMAD4 as a regulator of 4-hydroxytamoxifen (OHT) sensitivity in T47D cells. Clinical data analysis reveals downregulated SMAD4 expression in breast cancer tissues, correlating with poor prognosis. Following endocrine therapy, SMAD4 expression is further suppressed. Functional studies demonstrate that SMAD4 depletion induces endocrine resistance in vitro and in vivo by enhancing ER and ERBB signaling. Concomitant inhibition of ER and ERBB signaling leads to aberrant autophagy activation. Simultaneous inhibition of ER, ERBB, and autophagy pathways synergistically impacts SMAD4-depleted cells. Our findings unveil a mechanism whereby endocrine therapy-induced SMAD4 downregulation drives acquired resistance by integrating ER and ERBB signaling and suggest a rational treatment strategy for endocrine-resistant HR + HER2- breast cancer patients.

Integrated analysis of single-cell and bulk RNA sequencing data reveals the association between hypoxic tumor cells and exhausted T cells in predicting immune therapy response.

Continuous stimulation of tumor neoantigens and various cytokines in the tumor microenvironment leads to T cell dysfunction, but the specific mechanisms by which these key factors are distributed among different cell subpopulations and how they affect patient outcomes and treatment response are incompletely characterized. By integrating single-cell and bulk sequencing data of non-small cell lung cancer patients, we constructed a clinical outcome-associated T cell exhaustion signature. We discovered a significant association between the T cell exhaustion state and tumor cell hypoxia. Hypoxic malignant cells were significantly correlated with the proportion of exhausted T cells, and they co-occurred in patients at advanced stage. By analyzing the ligand-receptor interactions between these two cell states, we observed that T cells were recruited towards tumor cells through production of chemokines such as CXCL16-CXCR6 axis and CCL3/CCL4/CCL5-CCR5 axis. Based on 15 immune checkpoint blockade (ICB)-treatment cohorts, we constructed an interaction signature that can be used to predict the response to immune checkpoint blockade therapy. Among genes composed of the signature, CXCR6 alone has similarly high prediction efficacy (Area Under Curve (AUC) = 1, 0.89 and 0.73 for GSE126044, GSE135222 and GSE93157, respectively) with the signature and thus could serve as a potential biomarker for predicting immunotherapy response. Together, we have discovered and validated a significant association between exhausted T cells and hypoxic malignant cells, elucidating key interaction factors that significantly associated with response to immunotherapy.

Interaction between the gut microbiota and colonic enteroendocrine cells regulates host metabolism.

Nutrient handling is an essential function of the gastrointestinal tract. Hormonal responses of small intestinal enteroendocrine cells (EECs) have been extensively studied but much less is known about the role of colonic EECs in metabolic regulation. To address this core question, we investigated a mouse model deficient in colonic EECs. Here we show that colonic EEC deficiency leads to hyperphagia and obesity. Furthermore, colonic EEC deficiency results in altered microbiota composition and metabolism, which we found through antibiotic treatment, germ-free rederivation and transfer to germ-free recipients, to be both necessary and sufficient for the development of obesity. Moreover, studying stool and blood metabolomes, we show that differential glutamate production by intestinal microbiota corresponds to increased appetite and that colonic glutamate administration can directly increase food intake. These observations shed light on an unanticipated host-microbiota axis in the colon, part of a larger gut-brain axis, that regulates host metabolism and body weight.

Prophylactic efficacy of an intranasal spray with 2 synergetic antibodies neutralizing Omicron.

BACKGROUNDAs Omicron is prompted to replicate in the upper airway, neutralizing antibodies (NAbs) delivered through inhalation might inhibit early-stage infection in the respiratory tract. Thus, elucidating the prophylactic efficacy of NAbs via nasal spray addresses an important clinical need.METHODSThe applicable potential of a nasal spray cocktail containing 2 NAbs was characterized by testing its neutralizing potency, synergetic neutralizing mechanism, emergency protective and therapeutic efficacy in a hamster model, and pharmacokinetics/pharmacodynamic (PK/PD) in human nasal cavity.RESULTSThe 2 NAbs displayed broad neutralizing efficacy against Omicron, and they could structurally compensate each other in blocking the Spike-ACE2 interaction. When administrated through the intranasal mucosal route, this cocktail demonstrated profound efficacy in the emergency prevention in hamsters challenged with authentic Omicron BA.1. The investigator-initiated trial in healthy volunteers confirmed the safety and the PK/PD of the NAb cocktail delivered via nasal spray. Nasal samples from the participants receiving 4 administrations over a course of 16 hours demonstrated potent neutralization against Omicron BA.5 in an ex vivo pseudovirus neutralization assay.CONCLUSIONThese results demonstrate that the NAb cocktail nasal spray provides a good basis for clinical prophylactic efficacy against Omicron infections.TRIAL REGISTRATIONwww.chictr.org.cn, ChiCTR2200066525.FUNDINGThe National Science and Technology Major Project (2017ZX10202203), the National Key Research and Development Program of China (2018YFA0507100), Guangzhou National Laboratory (SRPG22-015), Lingang Laboratory (LG202101-01-07), Science and Technology Commission of Shanghai Municipality (YDZX20213100001556), and the Emergency Project from the Science & Technology Commission of Chongqing (cstc2021jscx-fyzxX0001).

DNA-scaffolded multivalent vaccine against SARS-CoV-2.

Short peptides are poor immunogens. One way to increase their immune responses is by arraying immunogens in multivalency. Simple and efficient scaffolds for spatial controlling the inter-antigen distance and enhancing immune activation are required. Here, we report a molecular vaccine design principle that maximally drives potent SARS-CoV-2 RBD subunit vaccine on DNA duplex to induce robust and efficacious immune responses in vivo. We expect that the DNA-peptide epitope platform represents a facile and generalizable strategy to enhance the immune response. STATEMENT OF SIGNIFICANCE: DNA scaffolds offer a biocompatible and convenient platform for arraying immunogens in multivalency antigenic peptides, and spatially control the inter-antigen distance. This can effectively enhance immune response. Peptide (instead of entire protein) vaccines are highly attractive. However, short peptides are poor immunogens. Our DNA scaffolded multivalent peptide immunogen system induced robust and efficacious immune response in vivo as demonstrated by the antigenic peptide against SARS-CoV-2. The present strategy could be readily generalized and adapted to prepare multivalent vaccines against other viruses or disease. Particularly, the different antigens could be integrated into one single vaccine and lead to super-vaccines that can protect the host from multiple different viruses or multiple variants of the same virus.

Real-world effectiveness of an intranasal spray A8G6 antibody cocktail in the post-exposure prophylaxis of COVID-19.

Previously, we identified an antibody combination A8G6 that showed promising efficacy in COVID-19 animal models and favorable safety profile in preclinical models as well as in a first-in-human trial. To evaluate the real-word efficacy of A8G6 neutralizing antibody nasal spray in post-exposure prophylaxis of COVID-19, an open-label, non-randomized, two-arm, blank-controlled, investigator-initiated trial was conducted in Chongqing, China (the register number: ChiCTR2200066416). High-risk healthy participants (18-65 years) within 72 h after close contact to COVID-19 patients were recruited and received a three-dose (1.4 mg/dose) A8G6 treatment daily or no treatment (blank control) for 7 consecutive days. SARS-CoV-2 infection occurred in 151/340 (44.4%) subjects in the blank control group and 12/173 (6.9%) subjects in the A8G6 treatment group. The prevention efficacy of the A8G6 treatment within 72 h exposure was calculated to be 84.4% (95% CI: 74.4-90.4%). Moreover, compared to the blank-control group, the time from the SARS-CoV-2 negative to the positive COVID-19 conversion was significantly longer in the AG86 treatment group (mean time: 3.4 days vs 2.6 days, p = 0.019). In the secondary end-point analysis, the A8G6 nasal treatment had no effects on the viral load at baseline SARS-CoV-2 RT-PCR positivity and the time of the negative COVID-19 conversion. Finally, except for 5 participants (3.1%) with general adverse effects, we did not observe any severe adverse effects related to the A8G6 treatment. In this study, the intranasal spray AG86 antibody cocktail showed potent efficacy for prevention of SARS-CoV-2 infection in close contacts of COVID-19 patients.

Time-restricted feeding alleviates metabolic implications of circadian disruption by regulating gut hormone release and brown fat activation.

Individuals with rotating and night shift work are highly susceptible to developing metabolic disorders such as obesity and diabetes. This is primarily attributed to disruptions in the circadian rhythms caused by activities and irregular eating habits. Time-restricted feeding (tRF) limits the daily eating schedules and has been demonstrated to markedly improve several metabolic disorders. Although an intricate relationship exists between tRF and circadian rhythms, the underlying specific mechanism remains elusive. We used a sleep disruption device for activity interference and established a model of circadian rhythm disorder in mice with different genetic backgrounds. We found that circadian rhythm disruption led to abnormal hormone secretion in the gut and elevated insulin resistance. tRF improved metabolic abnormalities caused by circadian rhythm disruption, primarily by restoring the gut hormone secretion rhythm and activating brown fat thermogenesis. The crucial function of brown fat in tRF was confirmed using a mouse model with brown fat removal. We demonstrated that chenodeoxycholic acid (CDCA) effectively improved circadian rhythm disruption-induced metabolic disorders by restoring brown fat activation. Our findings demonstrate the potential benefits of CDCA in reversing metabolic disadvantages associated with irregular circadian rhythms.

Augmenter of Liver Regeneration Monoclonal Antibody Promotes Apoptosis of Hepatocellular Carcinoma Cells.

Background and Aims: Hepatocellular carcinoma (HCC) is one of the most common types of cancer, often resulting in death. Augmenter of liver regeneration (ALR), a widely expressed multifunctional protein, has roles in liver disease. In our previous study, we reported that ALR knockdown inhibited cell proliferation and promoted cell death. However, there is no study on the roles of ALR in HCC. Methods: We used in vitro and in vivo models to investigate the effects of ALR in HCC as well as its mechanism of action. We produced and characterized a human ALR-specific monoclonal antibody (mAb) and investigated the effects of the mAb in HCC cells. Results: The purified ALR-specific mAb matched the predicted molecular weight of IgG heavy and light chains. Thereafter, we used the ALR-specific mAb as a therapeutic strategy to suppress tumor growth in nude mice. Additionally, we assessed the proliferation and viability of three HCC cell lines, Hep G2, Huh-7, and MHC97-H, treated with the ALR-specific mAb. Compared with controls, tumor growth was inhibited in mice treated with the ALR-specific mAb at 5 mg/kg, as shown by hematoxylin and eosin staining and terminal deoxynucleotidyl transferase dUTP nick end labeling. Simultaneous treatment with the ALR-specific mAb and adriamycin promoted apoptosis, whereas treatment with the ALR-specific mAb alone inhibited cell proliferation. Conclusions: The ALR-specific mAb might be a novel therapy for HCC by blocking extracellular ALR.

The Expression of LDL-R in CD8+ T Cells Serves as an Early Assessment Parameter for the Production of TCR-T Cells.

BACKGROUND/AIM: The quantity and the phenotypes of desired T cell receptor engineered T (TCR-T) cells in the final cell product determine their in vivo anti-tumor efficacy. Optimization of key steps in the TCR-T cell production process, such as T cell activation, has been shown to improve cell quality. MATERIALS AND METHODS: Using a modified TCR (mTCR) derived from mice transducing PBMCs, we assessed the proportions of low-density lipoprotein receptor (LDL-R) and mTCR expressing cells under the various activation conditions of CD3/CD28-Dynabeads or OKT3 via flow cytometry. RESULTS: We demonstrate that the proportion of T cells expressing LDL-R post activation is positively correlated with the percentage of mTCR+CD8+ T cells with their less differentiated subtypes in the final product. In addition, we show that shifting the CD3/CD28-Dynabeads activation duration from a typical 48 h to 24 h can significantly increase the production of the desired mTCR+CD8+ T cells. Importantly, the percentages of TCR-T cells with less-differentiated phenotypes, namely mTCR central memory T cells (TCM), were found to be preserved with markedly higher efficiency when T cell activation was optimized. CONCLUSION: Our findings suggest that the proportion of LDL-R+ T cells may serve as an early assessment parameter for evaluating TCR-T cell quality, possibly facilitating the functional and economical improvement of current adoptive therapy.

Bivalent intra-spike binding provides durability against emergent Omicron lineages: Results from a global consortium.

The SARS-CoV-2 Omicron variant of concern (VoC) and its sublineages contain 31-36 mutations in spike and escape neutralization by most therapeutic antibodies. In a pseudovirus neutralization assay, 66 of the nearly 400 candidate therapeutics in the Coronavirus Immunotherapeutic Consortium (CoVIC) panel neutralize Omicron and multiple Omicron sublineages. Among natural immunoglobulin Gs (IgGs), especially those in the receptor-binding domain (RBD)-2 epitope community, nearly all Omicron neutralizers recognize spike bivalently, with both antigen-binding fragments (Fabs) simultaneously engaging adjacent RBDs on the same spike. Most IgGs that do not neutralize Omicron bind either entirely monovalently or have some (22%-50%) monovalent occupancy. Cleavage of bivalent-binding IgGs to Fabs abolishes neutralization and binding affinity, with disproportionate loss of activity against Omicron pseudovirus and spike. These results suggest that VoC-resistant antibodies overcome mutagenic substitution via avidity. Hence, vaccine strategies targeting future SARS-CoV-2 variants should consider epitope display with spacing and organization identical to trimeric spike.

The binding affinity of a soluble TCR-Fc fusion protein is significantly improved by crosslinkage with an anti-Cß antibody.

The identification and cloning of tumor antigen-specific T cell receptors (TCRs) and the production of the soluble form of the TCR (sTCR) contributed to the development of diagnostic and therapeutic tools for cancer. Recently, several groups have reported the development of technologies for the production of sTCRs. The native sTCR has a very low binding affinity for the antigenic peptide/MHC (p/MHC) complex. In this study, we established a technology to produce high affinity, functional sTCRs. We generated a novel sTCR-Fc fusion protein composed of the TCR V and C regions of the TCR linked to the immunoglobulin (Ig) Fc region. A Western blot analysis revealed that the molecular weight of the fusion protein was approximately 60 kDa under reducing conditions and approximately 100-200 kDa under non-reducing conditions. ELISAs using various antibodies showed that the structure of each domain of the TCR-Fc protein was intact. The TCR-Fc protein immobilized by an anti-Cß antibody effectively bound to a p/MHC tetramer. An SPR analysis showed that the TCR-Fc protein had a low binding affinity (KD; 1.1 × 10(-5)M) to the p/MHC monomer. Interestingly, when the TCR-Fc protein was pre-incubated with an anti-Cß antibody, its binding affinity for p/MHC increased by 5-fold (2.2 × 10(-6)M). We demonstrated a novel method for constructing a functional soluble TCR using the Ig Fc region and showed that the binding affinity of the functional sTCR-Fc was markedly increased by an anti-Cß antibody, which is probably due to the stabilization of the Vα/Vß region of the TCR. These findings provide new insights into the binding of sTCRs to p/MHCs and will hopefully be instrumental in establishing functional sTCR as a diagnostic and therapeutic tool for cancer.

Rapid cloning of antigen-specific T-cell receptors by leveraging the cis activation of T cells.

It is commonly understood that T cells are activated via trans interactions between antigen-specific T-cell receptors (TCRs) and antigenic peptides presented on major histocompatibility complex (MHC) molecules on antigen-presenting cells. By analysing a large number of T cells at the single-cell level on a microwell array, we show that T-cell activation can occur via cis interactions (where TCRs on the T cell interact with the antigenic peptides presented on MHC class-I molecules on the same cell), and that such cis activation can be used to detect antigen-specific T cells and clone their TCR within 4 d. We used the detection-and-cloning system to clone a tumour-antigen-specific TCR from peripheral blood mononuclear cells of healthy donors. TCR cloning by leveraging the cis activation of T cells may facilitate the development of TCR-engineered T cells for cancer therapy.

Immune characteristics analysis reveals two key inflammatory factors correlated to the expressions of SARS-CoV-2 S1-specific antibodies.

The pandemic of COVID-19 caused by SARS-CoV-2 has made serious threats to the public health. Antibodies have been considered as promising therapeutics for the prevention and treatment of pathogens. So far, effectors that can influence the sustainability of SARS-CoV-2 specific antibodies in COVID-19 patients are still unclear. In this paper, we attempted to find potential key factors correlated with SARS-CoV-2 specific antibodies. Transcriptional analysis with the peripheral blood mononuclear cells (PBMCs) revealed proportional changes of immune cell subsets in COVID-19 convalescent patients, including a substantial decrease of monocytes and evident increase of dendritic cells (DCs). Moreover, we found that the gene expressions of chemokines associated with monocyte/macrophage were significantly up-regulated during the COVID-19 recovery phase. Most importantly, we found a set of 27 immune genes corresponding to a comparatively lower amount of SARS-CoV-2 specific antibodies, and identified two hub genes, IL1ß and IL6, the protein expressions of which exhibited negative correlation with the immunoglobulin G (IgG) levels in COVID-19 convalescent sera. In addition, we found that high expressions of these 2 hub genes during the convalescent stage were negatively associated with the plasma cell marker CD138. Our study presented two key inflammatory factors correlated to the low level of SARS-CoV-2 specific antibodies, which indicated the potential regulatory process of plasmatic antibodies levels in some COVID-19 convalescent patients.

CRISPR-based knockout screening identifies the loss of MIEF2 to enhance oxaliplatin resistance in colorectal cancer through inhibiting the mitochondrial apoptosis pathway.

The first-line anticancer agent oxaliplatin (OXL) is the preferred drug for treating colorectal cancer (CRC); however, the development of drug resistance is common in patients treated with OXL, which considerably reduces the efficacy of OXL-based regimens. By performing genome-wide CRISPR/Cas9 library knockdown screening, we found that mitochondrial elongation factor 2 (MIEF2) was among the top candidate genes. The OXL-resistant cell lines and organoids developed in the present study showed stable but low expression of MIEF2. Reduced MIEF2 expression may enhance CRC resistance to OXL by reducing mitochondrial stability and inhibiting apoptosis by decreasing cytochrome C release. In conclusion, among the different biomarkers of OXL resistance in CRC, MIEF2 may serve as a specific biomarker of OXL responsiveness and a potential target for the development of therapies to improve chemotherapeutic effectiveness.