Tonic signaling of chimeric antigen receptor (CAR), i.e., the spontaneous CAR activation in the absence of tumor antigen stimulation, is considered to be a pivotal event controlling CAR-T efficacy. However, the molecular mechanism underlying the spontaneous CAR signals remains elusive. Here, we unveil that positively charged patches (PCPs) on the surface of the CAR antigen-binding domain mediate CAR clustering and result in CAR tonic signaling. For CARs with high tonic signaling (e.g., GD2.CAR and CSPG4.CAR), reducing PCPs on CARs or boosting ionic strength in the culture medium during ex vivo CAR-T cell expansion minimizes spontaneous CAR activation and alleviates CAR-T cell exhaustion. In contrast, introducing PCPs into the CAR with weak tonic signaling, such as CD19.CAR, results in improved in vivo persistence and superior antitumor function. These results demonstrate that CAR tonic signaling is induced and maintained by PCP-mediated CAR clustering. Notably, the mutations we generated to alter the PCPs maintain the antigen-binding affinity and specificity of the CAR. Therefore, our findings suggest that the rational tuning of PCPs to optimize tonic signaling and in vivo fitness of CAR-T cells is a promising design strategy for the next-generation CAR.

Since B7-H3 is overexpressed or amplified in many types of solid tumors with a restricted expression in the normal tissues, it has been an emerging immunotherapeutic target for solid tumors. This review will focus on the structural designs of developing chimeric antigen receptors (CARs) targeting B7-H3. The expression, receptor, and function of the B7-H3, as well as a short overview of B7-H3-targeted monoclonal antibody therapy, are discussed. Finally, a detailed summary of B7-H3 redirected CAR-T and CAR-NK cell approaches utilized in preclinical models and currently ongoing or completed clinical trials are presented. It has been demonstrated that B7-H3-targeted CAR-based cell therapies were safe in initial trials, but their efficacy was limited. Employing the local delivery routes, the introduction of novel modifications promoting CAR-T persistence, and combined treatment with other standard therapies could improve the efficacy of B7-H3-targeted CAR-T cell therapy against solid tumors. Graphical Abstract Cancers develop by avoiding being discovered and destructed by the immune system. Cancer immunotherapy is characterized as an approach to assist the immune system to recognize and fight cancer by using uses some substances. Chimeric antigen receptor (CAR) is a protein designed to recognize a target of interest on tumor cells, immune cells armed with which are able to treat human cancers. B7-H3 is a type I transmembrane protein overexpressed in various solid tumors but scarcely detected in normal tissues. Moreover, it is associated with a poor prognosis for many cancer patients. Therefore, we discussed the role of B7-H3 in solid tumors, and concluded it as a promising therapeutic target for CAR-based immunotherapy. A number of B7-H3 CARs have been developed and tested in recent years, but which CAR construct targeting B7-H3 works best remains uncertain. Up till now, the design of B7-H3 CARs is still controversial and challenging. Hence, we summarized and compared the designs and efficacies of B7-H3 CARs utilized in preclinical models and currently ongoing or completed clinical trials in this review, and aimed to provide a better reference for future B7-H3 CAR design.

There is still no general method for discriminating between benign and malignant leukoplakia and identifying vocal fold leukoplakia. To evaluate the reliability of a morphological classification and the correlation between morphological types and pathological grades of vocal fold leukoplakia. A total of 375 patients with vocal fold leukoplakia between 2009 and 2015 were retrospectively reviewed. Two observers divided the vocal fold leukoplakia into flat and smooth, elevated and smooth, and rough type on the basis of morphological appearance. The inter-observer reliability was evaluated and the results of classification from both observers were compared with final pathological grades. Clinical characteristics between low risk and high risk group were also analyzed. The percentage inter-observer agreement of the morphological classification was 78.7% (κ=0.615, p<0.001). In the results from both observers, the morphological types were significantly correlated with the pathological grades (p₁<0.001, p₂<0.001, Kruskal-Wallis test; r₁=0.646, p₁<0.001, r₂=0.539, p₂<0.001, Spearman Correlation Analysis). Multivariate analysis showed patient's age (p=0.018), the size of lesion (p<0.001), and morphological type (p<0.001) were significantly different between low risk group and high risk group. Combined receiver operating characteristic curve analysis of significant parameters revealed an area under the receiver operating characteristic curve of 0.863 (95% CI 0.823-0.903, p<0.001). The proposed morphological classification of vocal fold leukoplakia was consistent between observers and morphological types correlated with pathological grades. Patient's age, the size of lesion, and morphological type might enable risk stratification and provide treatment guidelines for vocal fold leukoplakia.