Exploring the therapeutic potential of precision T-Cell Receptors (TCRs) in targeting KRAS G12D cancer through in vitro development

Objectives:The Kirsten rat sarcoma virus(KRAS)G12D oncogenic mutation poses a significant challenge in treating solid tumors due to the lack of specific and effective therapeutic interventions.This study aims to explore innovative approaches in T cell receptor(TCR)engineering and characterization to target the KRAS G12D7-16 mutation,providing potential strategies for overcoming this therapeutic challenge.Methods:In this innovative study,we engineered and characterized two T cell receptors(TCRs),KDA11-01 and KDA11-02 with high affinity for the KRAS G12D7-16 mutation.These TCRs were isolated from tumor-infiltrating lymphocytes(TILs)derived from tumor tissues of patients with the KRAS G12D mutation.We assessed their specificity and anti-tumor activity in vitro using various cancer cell lines.Results:KDA11-01 and KDA11-02 demonstrated exceptional specificity for the HLA-A*11:01-restricted KRAS G12D7-16 epitope,significantly inducing IFN-γrelease and eliminating tumor cells without cross-reactivity or alloreactivity.Conclusions:The successful development of KDA11-01 and KDA11-02 introduces a novel and precise TCR-based therapeutic strategy against KRAS G12D mutation,showing potential for significant advancements in cancer immunotherapy.

Role of triggering receptor expressed on myeloid cells 1/2 in secondary injury after cerebral hemorrhage

Intracerebral hemorrhage(ICH)is a common severe emergency in neurosurgery,causing tremendous economic pressure on families and society and devastating effects on patients both physically and psychologically,especially among patients with poor functional outcomes.ICH is often accompanied by decreased consciousness and limb dysfunction.This seriously affects patients’ability to live independently.Although rapid advances in neurosurgery have greatly improved patient survival,there remains insufficient evidence that surgical treatment significantly improves long-term outcomes.With in-depth pathophysiological studies after ICH,increasing evidence has shown that secondary injury after ICH is related to long-term prognosis and that the key to secondary injury is various immune-mediated neuroinflammatory reactions after ICH.In basic and clinical studies of various systemic inflammatory diseases,triggering receptor expressed on myeloid cells 1/2(TREM-1/2),and the TREM receptor family is closely related to the inflammatory response.Various inflammatory diseases can be upregulated and downregulated through receptor intervention.How the TREM receptor functions after ICH,the types of results from intervention,and whether the outcomes can improve secondary brain injury and the long-term prognosis of patients are unknown.An analysis of relevant research results from basic and clinical trials revealed that the inhibition of TREM-1 and the activation of TREM-2 can alleviate the neuroinflammatory immune response,significantly improve the long-term prognosis of neurological function in patients with cerebral hemorrhage,and thus improve the ability of patients to live independently.

Distribution of natural killer cell receptors in HIV infected individuals

Natural killer （NK） cells are bone marrow derived, large granular lymphocytes, comprising approximately 10% to 20% of the mononuclear cell fraction in normal peripheral blood. They form a part of the first line defense mechanism against tumoural and viral spreading^1-4 Unlike T and B cells, NK cells do not require gene rearrangement for assembly of their receptor genes; rather, NK cells discriminate potential target cells based on the levels of self major histocompatibility complex （MHC） class I expression on such cells^5.6 There are two kinds of NK cell receptors^2.7.8 Inhibitory receptors recognize MHC class I molecules and deliver a downregulatory signal that inactivates the lytic machinery of NK cells. Stimulatory receptors expressed by NK cells deliver an activation signal.

αβ T cell receptors as predictors of health and disease

The diversity of antigen receptors and the specificity it underlies are the hallmarks of the cellular arm of the adaptive immune system. T and B lymphocytes are indeed truly unique in their ability to generate receptors capable of recognizing virtually any pathogen. It has been known for several decades that T lymphocytes recognize short peptides derived from degraded proteins presented by major histocompatibility complex （MHC） molecules at the cell surface. Interaction between peptide-MHC （pMHC） and the T cell receptor （TCR） is central to both thymic selection and peripheral antigen recognition. It is widely assumed that TCR diversity is required, or at least highly desirable, to provide sufficient immune coverage. However, a number of immune responses are associated with the selection of predictable, narrow, or skewed repertoires and public TCR chains. Here, we summarize the current knowledge on the formation of the TCR repertoire and its maintenance in health and disease. We also outline the various molecular mechanisms that govern the composition of the pre-selection, naive and antigen-specific TCR repertoires. Finally, we suggest that with the development of high-throughput sequencing, common TCR ＇signatures＇ raised against specific antigens could provide important diagnostic biomarkers and surrogate predictors of disease onset, progression and outcome.

RELATIONSHIP BETWEEN SOMATOSTATIN RECEPTORS AND ACTIVATION OF HEPATIC STELLATE CELL

Objective To investigate the relationship between expression of somatostatin receptors(SSTRs) and activation of rat hepatic stellate cell (HSC). Methods HSCs were isolated from rats by in situ perfusion and single-step density gradient centrifugation, and then SSTR1-5 mRNA levels in the differentiated first passage HSCs were detected by means of reverse transcription polymerase chain reaction. On the other hand, hepatic fibrosis was induced in adult male Sprague-Dawley rats by carbon tetrachloride intoxication, and the expression of SSTR1-5 in normal as well as fibrotic liver was measured by immunohistochemical staining. Results SSTR mR-NA and SSTR could not be found in freshly isolated rat HSCs and normal rat liver. But SSTR1-3 mRNA appeared as HSCs became wholly activated, and SSTR1-3 could also be identified on the membrane of activated HSCs in the peri-sinusoid space, fibrous septa, etc. Conclusion The expression of SSTR1-3 in the rat HSC is closely related to its activation. This may reflect one of the main negative regulation mechanisms in the course of HSC activation.

P2X receptors in maintenance and differentiation of neural progenitor cells

Purinergic receptors are among the first cell surface receptors expressed during embryonic development （Burnstock and Ulrich, 2011）. These are characterized based on their pharmacological properties of being activated by adenosine or purine/pyrimidine nucleotides as P1 and P2 receptors. P2 receptors are further classified by their structure as P2Y metabotropic and P2X ionotropic receptors.

T_H1 AND T_H2 CELL ANTIGEN RECEPTORS IN EXPERIMENTAL LEISHMANIASIS

The complexity and ehronicity of parasitic infections have obscured the identification of biologically relevant antigens.Analysis of the T cell receptor repertoire used by mice infected with leishmania major revealed the expansion of a restricted population of CD4+ cells.These cells expressed the Vα 8-JαTA72,Vβ4 heterodimer in both progressive infection and protective immunity and across several major histocompatibility haplotypes.Thus,the same immunodominant parasite epitope drives the disparate outcomes of this infectious process, suggesting that candidate vaccine antigens selected by screening of immune individuals may be capable of exacerbating disease in genetically susceptible individuals.

Neurotrophins and their receptors in satellite glial cells following nerve injury

Peripheral neuropathy is a condition where damage resulting from mechanical or pathological mechanisms is inflicted on nerves within the peripheral nervous system （PNS）. Physical injury is the most common cause and may result in nerves being partially or completely severed, crushed, compressed or stretched. Other causes include metabolic or endocrine disorders, with e.g.,

Regulation of neural stem/progenitor cell functions by P2X and P2Y receptors

Neural stem/progenitor cells：Radial glial cells constitute multipotent cells in the ventricular zone,lining the wall of the lateral ventricle of the embryonic brain.They have the capacity to give rise to cells belonging to all three major linages（neurons,astrocytes and oligodendrocytes）of the nervous system（Tang and Illes,2017）.

The Roles and Mechanisms of TRAT1 in the Progression of Non-Small Cell Lung Cancer

Objective T cell receptor-associated transmembrane adaptor 1(TRAT1)is one of the hub genes regulating T cell receptors(TCRs).Herein,the roles of TRAT1 in the prognosis and immune microenvironment of non-small cell lung cancer(NSCLC)were investigated.Methods The expression and prognosis values of TRAT1 in NSCLC,and the relationship between TRAT1 expression levels and cancer immune cell infiltration was identified via the TIMER,UALCAN,TISIDB,and other databases.The mechanism of TRAT1 in NSCLC was analyzed using gene set enrichment analysis(GSEA).Results The expression level of TRAT1 was decreased in NSCLC tissues.Low TRAT1 expression was associated with shorter overall survival of patients with NSCLC and was related to gender,smoking,and tumor grade.TRAT1 was involved in regulating immune response,TCR signaling pathway,PI3K/AKT,and other processes.TRAT1 expression levels were positively correlated with immune cell infiltration in NSCLC.Conclusion Down-regulation of TRAT1 expression was associated with an unfavorable prognosis and immune infiltration of NSCLC.

Neurotoxic role of interleukin-17 in neural stem cell differentiation after intracerebral hemorrhage

Interleukin 17(IL-17)and its main producer,T cell receptorγδcells,have neurotoxic effects in the pathogenesis of intracerebral hemorrhage(ICH),aggravating brain injuries.To investigate the correlation between IL-17 and ICH,we dynamically screened serum IL-17 concentrations using enzyme-linked immunosorbent assay and explored the clinical values of IL-17 in ICH patients.There was a significant negative correlation between serum IL-17 level and neurological recovery status in ICH patients(r=–0.498,P<0.01).To study the neurotoxic role of IL-17,C57 BL/6 mice were used to establish an ICH model by injecting autologous blood into the caudate nucleus.Subsequently,the mice were treated with mouse neural stem cells(NSCs)and/or IL-17 neutralizing antibody for 72 hours.Flow cytometry,brain water content detection,Nissl staining,and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling results indicated that NSC transplantation significantly reduced IL-17 expression in peri-hematoma tissue,but there was no difference in T cell receptorγδcells.Compared with the ICH group,there were fewer apoptotic bodies and more Nissl bodies in the ICH+NSC group and the ICH+NSC+IL-17 group.To investigate the potential effect of IL-17 on directional differentiation of NSCs,we cultured mouse NSCs(NE-4 C)alone or co-cultured them with T cell receptorγδcells,which were isolated from mouse peripheral blood mononuclear cells,for 7 days.The results of western blot assays revealed that IL-17 secreted by T cell receptorγδcells reduced the differentiation of NSCs into astrocytes and neurons,while IL-17 neutralization relieved the inhibition of directional differentiation into astrocytes rather than neurons.In conclusion,serum IL-17 levels were elevated in the early stage of ICH and were negatively correlated with outcome in ICH patients.Animal experiments and cytological investigations therefore demonstrated that IL-17 probably has neurotoxic roles in ICH because of its inhibitory effects on the directional differentiation of NSCs.The application of IL-17 neutralizing antibody may promote the directional differentiation of NSCs into astrocytes.This study was approved by the Clinical Research Ethics Committee of Anhui Medical University of China(For human study:Approval No.20170135)in December 2016.All animal handling and experimentation were reviewed and approved by the Institutional Animal Care and Use Committee of Anhui Medical University(approval No.20180248)in December 2017.

Isolation and characteristics of autoreactive T cells specific to aggrecan G1 domain from rheumatoid arthritis patients

Our previous work showed that the cartilage proteoglycan aggrecan could induce an erosive polyarthritis and spondylitis in BALB/c mice and the GI globular domain of the aggrecan (GI) contained the arthritogenic region. To elucidate whether autoreactive T cells to G1 are expressed in rheumatoid arthritis patients, we analyzed the frequency of human G1-specific T cells in the peripheral blood of five rheumatoid arthritis patients and tried to establish G1-reactive T cell lines from these rheumatoid arthritis patients. The results showed that the G1-specific T cells in PBL were detectable at the range of 4.97 ±0.5 ×10-6 in peripheral blood lymphocytes. We have also generated 15 G1-specific T lymphocyte lines from these pateints with a standard split-well method. All these cells expressed fine specificity to human recombinant G1, but not to unrelated antigen. All the 15 lines expressed a panT cell marker and 13 of them selectively used the αβ T cell receptor. Two of them used rye T cell receptor. The 13 of these T cell lines was CD4 positive. One line expressed CD8. One line expressed both CD4 and CD8. Moreover, 14 out of 15 lines expressed the Th-1 cytokine profile, characterized by interferon-γpositivity and IL-4 negativity. No Th-2 type cell line was generated. These data provide strong evidence in favor of the presence of autoreactive T cells in the rheumatoid arthritis pateints. What is the mechanism(s) that these autoreactive T cells attack self-target and whether these G1-specific, Th-1 type T cell lines can induce arthritis in immune deficiency mice are currently under investigation.

Activation-induced cell death in B lymphocytes

Upon encountering the antigen (Ag), the immune system can either develop a specific immune response or enter a specific state of unresponsiveness, tolerance. The response of B cells to their specific Ag can be activation and proliferation, leading to the immune response, or anergy and activation-induced cell death (AICD), leading to tolerance. AICD in B lymphocytes is a highly regulated event initiated by crosslinking of the B cell receptor (BCR). BCR engagement initiates several signaling events such as activation of PLCr, Ras, and PI3K, which generally speaking, lead to survival. However, in the absence of survival signals (CD40 or IL-4R engagement), BCR crosslinking can also promote apoptotic signal transduction pathways such as activation of effector caspases, expression of pro-apoptotic genes, and inhibition of pro-survival genes. The complex interplay between survival and death signals determines the B cell fate and, consequently, the immune response.

B cell receptor signaling pathway involved in benign lymphoepithelial lesions of the lacrimal gland

AIM：To detect the expression of B cell receptor signaling pathway（BCRSP） in lacrimal gland benign lymphoepithelial lesions（LGBLEL）.METHODS：Gene microarray was used to compare whole-genome expression in lacrimal gland tissues from LGBLEL patients to tissues from orbital cavernous hemangioma（control tissues）. Expression of BCRSP was confirmed by polymerase chain reaction（PCR） and immunohistochemistry. RESULTS：The expression of 22 genes of the BCRSP increased significantly in LGBLEL patients. PCR analysis showed that CD22, CR2, and BTK were all highly expressed in LGBLEL tissues. Immunohistochemical analysis showed that CR2 protein was present in LGBLEL, but CD22 and BTK proteins were negative. CR2, CD22, and BTK were not observed in the orbital cavernous hemangiomas with either PCR or immunohistochemistry. CONCLUSION：BCRSP might be involved in the pathogenesis of LGBLEL.

Novel Association of Killer Cell Immunoglobulin-like Receptor Genes with EBV-infectious Diseases in Children

Killer cell immunoglobulin-like receptors （KIRs） which are mainly expressed on natural killer （NK） cells are implicated in many virus infections. However, it is unclear whether or not KIRs are associated with susceptibility to Epstein-Barr virus （EBV） infection related diseases. Therefore, the purpose of our study was to investigate possible correlation between polymorphisms of KIR genes and infectious mononucleosis （IM）/EBV-associated hemophagocytic Iymphohistiocytosis （EBV-HLH）. The polymorphisms of KIR genes were detected by polymerase chain reaction with sequence-specific primers （PCR-SSP）. The results would contribute to clarify the association of KIRs with EBV induced diseases, and provide new insights into the role of NK cells and innate immune response against viral infections and/or subsequent progression.

Detection of Minimal Leukemic Cells in Cerebral Spinal Fluid of Children with Acute Lymphoblastic Leukemia Using the Polymerase Chain Reaction Technique

Vδ2Dδ3 rearrangements of T cell receptor (TCR) gene from cerebralspinal fluid (CSF) cells was detected for diagnosis and monitoring of central nervous system leukemia (CNSL) in children with acute lymphoblastic leukemia (ALL) using polymerase chain reaction (PCR) technique. 20 patients were studied and in 12 of them the results of PCR and dot hybridization with clonospecific probes were positive, showing the presence of minimal blast cells in CSF. Our study suggested that the PCR method is an effective tool for clinical diagnosis of CNSL and is much more sensitive than routine CSF examination.

CAR T Cell Therapy for Hematological Malignancies

As a rapidly progressing field in oncology,the adoptive transfer of T cells that have been genetically modified with chimeric antigen receptors(CARs)has shown striking efficacy in the management of hematological malignancies and has been reported in a number of clinical trials.of note,CAR T cell therapy has shown extraordinary potential,especially in relapsed/refractory patients.However,there are still challenges regarding the further development of this strategy,spanning from engineering and manufacturing issues,to limited applications,to accompanying toxicities.In this review,we will summarize the general knowledge of this novel method,including receptor composition,applications,adverse events and challenges.Additionally,we will propose several comprehensive recommendations.

Bone marrow mesenchymal stem cells transplantation promotes the release of endogenous erythropoietin after ischemic stroke

This study investigated whether bone marrow mesenchymal stem cell（BMSC） transplantation protected ischemic cerebral injury by stimulating endogenous erythropoietin. The model of ischemic stroke was established in rats through transient middle cerebral artery occlusion. Twenty-four hours later, 1 × 106 human BMSCs（h BMSCs） were injected into the tail vein. Fourteen days later, we found that h BMSCs promoted the release of endogenous erythropoietin in the ischemic region of rats. Simultaneously, 3 μg/d soluble erythropoietin receptor（s EPOR） was injected into the lateral ventricle, and on the next 13 consecutive days. s EPOR blocked the release of endogenous erythropoietin. The neurogenesis in the subventricular zone was less in the h BMSCs ＋ s EPOR group than in the h BMSCs ＋ heat-denatured s EPOR group. The adhesive-removal test result and the modified Neurological Severity Scores（m NSS） were lower in the h BMSCs ＋ s EPOR group than in the heat-denatured s EPOR group. The adhesive-removal test result and m NSS were similar between the h BMSCs ＋ heat-denatured s EPOR group and the h BMSCs ＋ s EPOR group. These findings confirm that BMSCs contribute to neurogenesis and improve neurological function by promoting the release of endogenous erythropoietin following ischemic stroke.

AduoLa Fuzhenglin Down-regulates Microwave-induced Expression of β_1-adrenergic Receptor and Muscarinic Type 2 Acetylcholine Receptor in Myocardial Cells of Rats

This paper is aimed to study the effect of ADL on expression of ~z-AR and Mz-AchR in myocardial cells of rats exposed to microwave radiation. Immunohistochemistry, Western blot and image analysis were used to detect the expression of ~I-AR and Mz-AchR in myocardial cells at 7 and 14 d after microwave exposure. The results show that the expression level was higher in microwave exposure group and 0.75 g/（kg.d） ADL group than in sham operation group and significantly lower in 1.5 and 3.0 g/（kg.d） ADL groups than in microwave group. So we have a conclusion that the expression of I^z-AR and Mz-AchR is down-regulated in myocardial cells of rats exposed to microwave radiation. ADL can protect rats against microwave-induced heart tissue injury.

Engineered T cells and their therapeutic applications in autoimmune diseases

Chimeric antigen receptor T cells(CAR-T cells) are engineered recombinant T cells, which were initially used to treat hematopoietic malignancies and are now widely used in the treatment of various diseases. Considering their intrinsic targeting efficiency, CAR-T cells show considerable potential in the treatment of autoimmune diseases.Furthermore, regulatory T cells(Treg), a subset of CD4 T cells exhibiting immunosuppressive functions,have attracted increasing attention regarding CARTreg cell production. In this review, we report on recent developments in preclinical and clinical studies on CAR-T cells in autoimmune diseases and provide an outlook on opportunities and challenges of CAR-T application in such diseases.