NY-ESO-1-specific T cell receptor-engineered T cells and Tranilast, a TRPV2 antagonist bivalent treatment enhances the killing of esophageal cancer: a dual-targeted cancer therapeutic route.

BACKGROUND: Esophageal cancer (EC) is a global canker notorious for causing high mortality due to its relentless incidence rate, convoluted with unyielding recurrence and metastasis. However, these intricacies of EC are associated with an immoderate expression of NY-ESO-1 antigen, presenting a lifeline for adoptive T cell therapy. We hypothesized that naturally isolated higher-affinity T cell receptors (TCRs) that bind to NY-ESO-1 would allow T lymphocytes to target EC with a pronounced antitumor response efficacy. Also, targeting TRPV2, which is associated with tumorigenesis in EC, creates an avenue for dual-targeted therapy. We exploited the dual-targeting antitumor efficacy against EC. METHODS: We isolated antigen-specific TCRs (asTCRs) from a naive library constructed with TCRs obtained from enriched cytotoxic T lymphocytes. The robustness of our asTCRs and their TCR-T cell derivatives, Tranilast (TRPV2 inhibitor), and their bivalent treatment were evaluated with prospective cross-reactive human-peptide variants and tumor cells. RESULTS: Our study demonstrated that our naive unenhanced asTCRs and their TCR-Ts perpetuated their cognate HLA-A*02:01/NY-ESO-1(157-165) specificity, killing varying EC cells with higher cytotoxicity compared to the known affinity-enhanced TCR (TCRe) and its wild-type (TCR0) which targets the same NY-ESO-1 antigen. Furthermore, the TCR-Ts and Tranilast bivalent treatment showed superior EC killing compared to any of their monovalent treatments of either TCR-T or Tranilast. CONCLUSION: Our findings suggest that dual-targeted immunotherapy may have a superior antitumor effect. Our study presents a technique to evolve novel, robust, timely therapeutic strategies and interventions for EC and other malignancies.

HIF-1α promotes kidney organoid vascularization and applications in disease modeling.

BACKGROUND: Kidney organoids derived from human pluripotent stem cells (HiPSCs) hold huge applications for drug screening, disease modeling, and cell transplanting therapy. However, these applications are limited since kidney organoid cannot maintain complete morphology and function like human kidney. Kidney organoids are not well differentiated since the core of the organoid lacked oxygen, nutrition, and vasculature, which creates essential niches. Hypoxia-inducible factor-1 α (HIF-1α) serves as a critical regulator in vascularization and cell survival under hypoxia environment. Less is known about the role of HIF-1α in kidney organoids in this regard. This study tried to investigate the effect of HIF-1α in kidney organoid vascularization and related disease modeling. METHODS: For the vascularization study, kidney organoids were generated from human induced pluripotent stem cells. We overexpressed HIF-1α via plasmid transfection or treated DMOG (Dimethyloxallyl Glycine, an agent for HIF-1α stabilization and accumulation) in kidney progenitor cells to detect the endothelium. For the disease modeling study, we treated kidney organoid with cisplatin under hypoxia environment, with additional HIF-1α transfection. RESULT: HIF-1α overexpression elicited kidney organoid vascularization. The endothelial cells and angiotool analysis parameters were increased in HIF-1α plasmid-transfected and DMOG-treated organoids. These angiogenesis processes were partially blocked by VEGFR inhibitors, semaxanib or axitinib. Cisplatin-induced kidney injury (Cleaved caspase 3) was protected by HIF-1α through the upregulation of CD31 and SOD2. CONCLUSION: We demonstrated that HIF-1α elicited the process of kidney organoid vascularization and protected against cisplatin-induced kidney organoid injury in hypoxia environment.

SYVN1 ubiquitinates FoxO1 to induce ß-catenin nuclear translocation, PD-L1-mediated metastasis, and immune evasion of hepatocellular carcinoma.

BACKGROUND: A high incidence of hepatocellular carcinoma (HCC), the most frequently diagnosed form of liver cancer, is observed in Africa and Asia. SYVN1 is upregulated in HCC; however, the biological roles of SYVN1 in immune evasion remain unclear. METHODS: RT-qPCR and western blot were employed to detect the expression levels of SYVN1 and the key molecules in HCC cells and tissues. Flow cytometry was used to determine the proportion of T cells, and an ELISA assay was used to determine the amount of IFN-γ secreted. Cell viability was monitored by CCK-8 and colony formation assays. The metastatic properties of HCC cells were detected by Transwell assays. Bioinformatics analysis, ChIP, and luciferase assays were used to study the transcriptional regulation of PD-L1. Co-IP was used to detect direct interaction between SYVN1 and FoxO1, as well as the ubiquitination of FoxO1. The in vitro findings were validated in xenograft and lung metastasis models. RESULTS: In HCC cells and tissues, SYVN1 was upregulated while FoxO1 was downregulated. SYVN1 knockdown or FoxO1 overexpression reduced PD-L1 expression, and inhibited immune evasion, cell growth, and metastasis in HCC cells. Mechanistically, FoxO1 regulated PD-L1 transcription in a ß-catenin-independent or -dependent manner. Functional studies further showed that SYVN1 promoted immune evasion, cell proliferation, migration and invasion via facilitating ubiquitin-proteasome-dependent degradation of FoxO1. In vivo investigations showed that silencing of SYVN1 inhibited immune evasion and metastasis of HCC cells, possible via the FoxO1/PD-L1 axis. CONCLUSION: SYVN1 regulates FoxO1 ubiquitination to stimulate ß-catenin nuclear translocation and promotes PD-L1-mediated metastasis and immune evasion in HCC.

Transcription factor EB-mediated mesenchymal stem cell therapy induces autophagy and alleviates spinocerebellar ataxia type 3 defects in neuronal cells model.

Defects in ataxin-3 proteins and CAG repeat expansions in its coding gene ATXN3 cause Spinocerebellar Ataxia Type 3 (SCA3) or Machado-Joseph disease (MJD) polyglutamine neurodegenerative disease. The mutant proteins aggregate as inclusion bodies in cells and compete with wild-type ataxin-3, which leads to neuronal dysfunction or death and impairs Beclin1-mediated autophagy. It has been reported that Mesenchymal stem cells (MSCs) can reliably treat several neurodegenerative diseases. Herein, we used a Transcription Factor EB (TFEB) nuclear translocation-mediated MSCs co-culture approach to reconstitute autophagy and lysosomal biogenesis, and reduce SCA3-like behaviors in induced pluripotent stem cells (iPSCs)-derived neuron cells models. Our iPSCs model showed enhanced expression of autophagy proteins, attenuated the expression and toxic effects of mutant ataxin-3 on neurons, and alleviated the effects of ataxin-3 on autophagy. Therefore, MSCs are associated with autophagy-inducing therapy and compared to animal models, our MSCs co-culture could be used as a novel and potential therapeutic approach to study SCA3 disease and other neurodegenerative diseases.

A rapid and sensitive CRISPR/Cas12a based lateral flow biosensor for the detection of Epstein-Barr virus.

Nasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in the world, and several studies have associated Epstein-Barr virus (EBV) with NPC occurrence and development. EBV-PCR (polymerase chain reaction), in situ hybridization and immunoassays are the most common methods for NPC identification. However, these approaches have drawbacks, which include tedious procedures and false results. Therefore, a rapid, accurate, and sensitive clinical diagnostic method for the prognosis of EBV-related diseases is needed. In this study, we developed a simple and sensitive approach for EBV detection based on the combination of CRISPR-Cas12a and a lateral flow biosensor (LFB). Cas12a exhibits collateral cleavage propensity of both target DNA and any single-stranded(ss) DNA in the vicinity (herein referred to as a reporter). The LFB test line contained an ssDNA probe complementary to the reporter. In the presence of the target, Cas12a trans-cleaved the ssDNA reporter, which resulted in the inability of cleaved sequences to bind the LFB test line. With a PCR pre-amplification of the target (45 min), the assay achieved a sensitivity of 7.1 × 10-14 M (∼42 000 copies per µl) both in plasmid and plasmid-spiked samples. The assay attained a high specificity in the presence of various bacteria and applicability in EBV Burkitt's lymphoma serum samples. This method could be applied for the detection of EBV and other infectious diseases.

Ultrasensitive Fluorometric Angling Determination of Staphylococcus aureus in Vitro and Fluorescence Imaging in Vivo Using Carbon Dots with Full-Color Emission.

Rapid, accurate, and safe screening of foodborne pathogenic bacteria is essential to effectively control and prevent outbreaks of foodborne illness. Fluorescent sensors constructed from carbon dots (CDs) and nanomaterial-based quenchers have provided an innovative method for screening of pathogenic bacteria. Herein, an ultrasensitive magnetic fluorescence aptasensor was designed for separation and detection of Staphylococcus aureus (S. aureus). Multicolor fluorescent CDs with a long fluorescent lifetime (6.73 ns) and high fluorescence stability were synthesized using a facile hydrothermal approach and modified cDNA as a highly sensitive fluorescent probe. CD fluorescence was quenched by Fe3O4 + aptamer via fluorescence resonance energy transfer (FRET). Under optimal conditions, the FRET-based aptasensor can detect S. aureus accompanied by a wide linear range of 50-107 CFU·mL-1 and a detection limit of 8 CFU·mL-1. Compared with other standard methods, this method was faster and more convenient, and the entire test was finished within 30 min. The capability of the aptasensor was simultaneously investigated on food samples. Additionally, the developed CDs exhibited excellent biocompatibility and were thus applied as fluorescent probes for bioimaging both in vitro and in vivo. This new platform provided an excellent application of the CDs for detecting and bioimaging pathogenic bacteria.