5-fluorouracil (5-Fu) is still recognized as the mainstay in colorectal cancer chemotherapy, but the response rate of 5-Fu in colorectal cancer is less than 50%. Our previous mRNA microarray data revealed that BRCC3, a component of the BRCA1-BRCA2-BRCC3 DNA repair complex, had a direct relationship with B7-H3, an immunoglobulin that is upregulated in tumor tissue and associated with metastasis and poor prognosis. Real-time PCR and western blot analysis confirmed that the expression of both BRCC3 mRNA and protein, respectively, were elevated following B7-H3 overexpression in SW480 cells; likewise, BRCC3 expression decreased after B7-H3 was knocked down in HCT-8 cells. DNA comet assay results indicate an inverse correlation between the extent of 5-Fu-induced DNA damage and the expression level of B7-H3 in both SW480- and HCT-8-based cell lines. In SW480 cells that overexpress B7-H3, knockdown of BRCC3 similarly permitted greater 5-Fu-induced DNA damage. Altogether, results suggest that BRCC3 may play a role in B7-H3-induced 5-Fu resistance, such that B7-H3 upregulates BRCC3 expression, enhancing DNA repair in colorectal cancer cells.

Previous studies have investigated the prognostic significance of B7 homolog 3 (B7-H3) in non-small cell lung cancer (NSCLC), however, the results remain controversial. This study was aimed to determine the correlation between B7-H3 and survival as well as clnicalpathological characteristics in NSCLC using meta-analysis. We searched the electronic databases of PubMed, Embase, Web of Science, and China National Knowledge Infrastructure (CNKI) for relevant studies up to October 9, 2016. Pooled hazard ratios (HRs) and 95% confidence intervals (CIs) were used to estimate the impact of B7-H3 on overall survival (OS). Combined odds ratios (ORs) and 95%CIs were utilized to evaluate the correlations between B7-H3 and clinicalpathological features. This meta-analysis finally included 7 studies with 864 patients. The results showed that B7-H3 had no significant association with OS (HR=0.88, 95%CI: 0.36-2.13, p=0.776). High B7-H3 expression was a significant indicator of lymph node metastasis (OR=3.92, 95%CI: 2.65-5.81, p<0.001), and advanced TNM stage (OR=3.53, 95%CI: 2.45-5.09, p<0.001). B7-H3 has the potential to serve as a marker of tumor aggressiveness and lymph node metastasis in NSCLC. However, due to several limitations, further large-scale studies are needed to validate our results.

Central nervous system (CNS) is an increasingly common site of isolated metastasis for patients with Stage 4 neuroblastoma. To explore the microRNA (miRNA) profile of this metastatic process, miRNA sequencing was performed to identify miRNA sequence families with differential expression between tumor pairs (pre-CNS primary and CNS metastasis) from 13 patients with Stage 4 neuroblastoma. Seven miRNA sequence families had distinct expression in CNS metastases when compared with their corresponding pre-CNS primaries. MiR-7 was upregulated (3.75-fold), and miR-21, miR-22, miR-29a, miR-143, miR-199a-1-3p, and miR-199a-1-5p were downregulated (3.5-6.1-fold), all confirmed by quantitative reverse transcription-PCR. MiR-29a, previously shown to be downregulated in a broad spectrum of solid tumors including neuroblastoma, had the most significant decrease in all 13 CNS metastases (P = 0.001). Its known onco-targets CDC6, CDK6, and DNMT3A, as well as B7-H3, an inhibitory ligand for T cells, and natural killer cells, were found to have higher differential expression in these 13 CNS metastases when compared with their paired primaries. Additionally, miR-29a expression in primary tumors was significantly lower among patients who eventually relapsed in the CNS. Irrespective of the amplification status of MYCN, which is known to be associated with metastasis, pre-CNS primaries, and CNS metastases had significantly lower miR-29a expression than non-CNS primary tumors. Among MYCN amplified cell lines, those from CNS relapse also had lower miR-29a expression than non-CNS relapse. These findings raised the hypothesis that miR-29a could be a biomarker for neuroblastoma CNS metastasis, and its downregulation may play a pivotal role in CNS progression.

B7-H3, a co-stimulatory molecule, has been found expressed in ovarian cancer, but its role and mechanism is not clear. In this study, we further verified the expression of B7-H3 in ovarian carcinoma and normal epithelial ovarian tissues. Three ovarian cancer cell lines, A2780, SKOV3 and HO8910 were selected to explore the effects of B7-H3 on proliferation, apoptosis, migration and invasion. We found that B7-H3 was mainly located in the cytoplasm of ovarian cancer cells as determined by immunofluorescence staining. The ability of cell invasion, migration, proliferation decreased after silencing B7-H3 whereas the apoptosis increased, which was related to the upregulation of Bax, caspase-8, cleaved caspase-8 and the downregulation of Bcl-2, Bcl-xl, matrix metalloproteinase-2 (MMP2) by western blotting. In addition, B7-H3 enhanced the H08910 cell capacities in invasion, migration and proliferation. Expression of the phosphorylation signal transducer and activator of transcription 3 (pStat3) molecules and their upstream molecules phosphorylation Janus kinase 2 (pJak2) were significantly increased. In order to investigate whether B7-H3 plays a role in this pathway, we treated the overexpressed HO8910 cells with AG490 (inhibitors of Jak2). Our findings revealed that B7-H3 affect ovarian cancer progression through the Jak2/Stat3 pathway, indicating that B7-H3 has the potential to be a useful prognostic marker.

The success of immune checkpoint inhibitors (ICIs), notably anti-cytotoxic T lymphocyte associated antigen-4 (CTLA-4) as well as inhibitors of CTLA-4, programmed death 1 (PD-1), and programmed death ligand-1 (PD-L1), has revolutionized treatment options for solid tumors. However, the lack of response to treatment, in terms of de novo or acquired resistance, and immune related adverse events (IRAE) remain as hurdles. One mechanisms to overcome the limitations of ICIs is to target other immune checkpoints associated with tumor microenvironment. Immune checkpoints such as lymphocyte activation gene-3 (LAG-3), T cell immunoglobulin and ITIM domain (TIGIT), T cell immunoglobulin and mucin-domain containing-3 (TIM-3), V-domain immunoglobulin suppressor of T cell activation (VISTA), B7 homolog 3 protein (B7-H3), inducible T cell costimulatory (ICOS), and B and T lymphocyte attenuator (BTLA) are feasible and promising options for treating solid tumors, and clinical trials are currently under active investigation. This review aims to summarize the clinical aspects of the immune checkpoints and introduce novel agents targeting these checkpoints.

Keywords: B7-H3; BTLA; ICOS; LAG-3; TIGIT; TIM-3; VISTA; immune checkpoint.

Whereas B7-1/B7-2 and CD28/cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) serve as the main switches regulating the clonal composition of activated naive T cells, other B7 family members fine-tune the expansion and properties of activated T cells. Inducible costimulatory molecule (ICOS)-B7h promotes T-dependent antibody isotype switching and expansion of effector cells. Effector T cells trafficking into inflamed tissues interact with antigen-presenting cells there and are regulated by PD-1 and its ligands. B7-H3 and B7x could control the interaction between effector T cells and the peripheral tissues. The different varieties of regulatory T cells could regulate both naive T cell activation and effector function through costimulatory receptor/ligands.

No treatment prolongs the survival of malignant mesothelioma (MM) patients. Since MM elicits anti-tumor host's immune responses, immunotherapy represents a promising strategy for its control. Immunomodulatory antibodies against components of the B7 family of immunomodulatory molecules that regulate T cell activation are being investigated in human malignancies including MM. The expression of B7-H3, a new component of the B7 family was investigated in primary cultures of human mesothelial cells (HMC) and in MM cell lines by flow cytometry and molecular analyses, and in MM tissues by immunohistochemistry. The role of DNA hypomethylating agents in modulating levels of B7-H3 expression in MM cells was also studied. Reverse transcriptase-polymerase chain reaction (RT-PCR) demonstrated that B7-H3 mRNA was consistently detectable in mesothelial and MM cells investigated; however, real-time quantitative RT-PCR analyses showed highly heterogeneous levels of B7-H3 mRNA among investigated MM cells. The analysis of B7-H3 protein expression indicated that comparable levels of B7-H3 were expressed on both cell types. Treatment with the DNA hypomethylating agent 5-aza-2'-deoxycytidine did not significantly affect the expression of B7-H3 mRNA in MM cells. In vivo, while B7-H3 was expressed in all 13 tumor biopsies of the epithelial variant, with high levels in 54% of cases, it was rarely detectable in spindle type MM in which 1/5 biopsies weakly expressed B7-H3. These findings suggest that B7-H3 is a promising target for new immunotherapeutic strategies in MM, with particular emphasis in the epithelial variant.

In mammals, professional APCs induce adaptive immunity via the activation of T cells. During this process, B7 family molecules present upon APCs are known to play crucial roles in optimal T cell stimulation. In contrast, the confirmation of APCs in a nonmammalian vertebrate has yet to be achieved. To obtain further insights into the evolutionary origin of APCs, we have identified three members of the B7 family in the teleost Takifugu rubripes (fugu): B7-H1/DC, B7-H3, and B7-H4. The three fugu B7s were expressed on the surface of blood monocytes. The B7(+) monocytes, which are composed of at least two distinct populations, expressed the MHC class II component gene. The fugu B7 molecules bound to activated T cells, indicating that putative B7 receptors were expressed upon T cells. Fugu B7-H1/DC inhibited T cell proliferation concomitant with increasing levels of both IL-10 and IFN-gamma expression, whereas both B7-H3 and B7-H4 promoted T cell growth following IL-2 induction and the suppression of IL-10. These observations indicate that fugu B7s regulate T cell responses via receptors upon T cells. We suggest that fish B7(+) monocytes are APCs and that a costimulatory system has already developed in fish via the evolutionary process.

OBJECTIVE

Negative T-cell costimulatory pathways regulate T-cell responses and are of great interest in the development of tolerance-inducing strategies in transplantation. This article provides an update of major contributions to the understanding of the known negative costimulatory pathways, and reviews recent studies on the effects of targeting of these pathways in alloimmunity.

RESULTS

Graft tissue expression of negative costimulatory molecules, especially programmed death receptor ligand 1, plays a major role in regulation of alloimmune responses. The recently demonstrated programmed death receptor ligand 1:B7-1 interaction highlights the complexity and possible redundancy/hierarchy of the functions of these pathways in vivo. CD160 was identified as a ligand of Herpes virus entry mediator and has been established as a new coinhibitory molecule. A soluble form of B7-H3 with possible functional roles has been identified in mice and humans.

CONCLUSIONS

The understanding of negative costimulatory pathways is embarrassed by complex interactions between simultaneously activated positive and negative costimulatory pathways among themselves and with immunosuppressive agents, differential expression of these molecules on different immune cell subsets as well as their expression in parenchymal cells of transplanted tissues, all of them clearly affecting their functions. Further elucidation of these novel concepts is pertinent for targeting these pathways in translational studies in near future.

B7-H3 (CD276), a member of the B7 superfamily, is an important factor in downregulating immune responses against tumors. It is also aberrantly expressed in many human malignancies. Beyond immune regulatory roles, its overexpression has been linked to invasive metastatic potential and poor prognosis in patients with cancer. Antibody-dependent cell-mediated cytotoxicity strategies targeting B7-H3 are currently in development, and early-phase clinical trials have shown encouraging preliminary results. To understand the role of B7-H3 in pediatric central nervous system (CNS) malignancies, a comprehensive panel of primary CNS tumors of childhood was examined by immunohistochemistry for levels and extent of B7-H3 expression. In addition, B7-H3 m-RNA expression status and association with overall survival in various pediatric CNS tumor types was accessed by curating publicly available patient gene expression data sets derived from bioinformatics analysis and visualization platforms (GlioVis). We demonstrate that B7-H3 is broadly expressed in pediatric glial and nonglial CNS tumors, and its aberrant expression, as determined by immunohistochemical staining intensity, correlates with tumor grade. Moreover, high B7-H3 m-RNA expression is significantly associated with worse survival and could potentially improve prognostication in various brain tumor types of childhood. B7-H3 can be used as a therapeutic target, given its tumor selectivity and the availability of targeted therapeutic agents to this antigen.

Biomarkers that can serve as diagnostic and prognostic indicators of esophageal squamous cell carcinoma (ESCC) are urgently needed to help improve patient outcomes. Here, the expression of epidermal growth factor receptor (EGFR) and costimulatory molecule B7-H3, both of which have been implicated in tumor onset and progression in certain tumors, was investigated in relation to the clinical characteristics and survival outcomes of patients with ESCC. ESCC tissue samples were analyzed for 100 patients. Tumor and patient characteristics were recorded. Tissues were investigated for EGFR and B7-H3 staining by immunohistochemistry. Patients were followed for up to 96 months to determine overall survival (OS) and progression-free survival (PFS). High expression for EGFR (68.0%) and B7-H3 (66.0%) was observed in the majority of cases. High expression of either EGFR or B7-H3 was correlated with tumor invasion depth and clinical stage (P<0.05). Further, high expression of either EGFR or B7-H3 was correlated with worse survival outcomes. The estimated OS (38.1 months) and PFS (13.4 months) of patients with high expression of EGFR were lower than those of patients with low expression (69.3 and 68.1 months, P<0.05). The estimated OS (31.1 months) and PFS (13.1 months) of patients with high expression of B7-H3 were also lower than those of patients with low expression (69.3 and 66.6 months, P<0.05). Indeed, Cox multiple regression showed that OS and PFS were correlated with EGFR (relative risk =1.853, 1.875, respectively) and B7-H3 (relative risk =1.886, 2.061, respectively) (all P<0.05) expression level. Thus, EGFR and B7-H3 are highly expressed in tumor tissues of patients with ESCC. Their expression levels are correlated with tumor severity and survival, and therefore these may be viable biomarkers to aid in prognosis determination.

Background: Immunotherapy with CAR T-cells is actively being explored for pediatric brain tumors in preclinical models and early phase clinical studies. At present it is unclear which CAR target antigens are consistently expressed across different pediatric brain tumor types. In addition, the extent of HLA class-I expression is unknown, which is critical for tumor recognition by conventional αβTCR T-cells.

Methods: We profiled 49 low- and high-grade pediatric brain tumor patient-derived orthotopic xenografts (PDOX) by flow analysis for the expression of five CAR targets (B7-H3, GD2, IL13Rα2, EphA2, HER2), and HLA class-I. In addition, we generated B7-H3-CAR T-cells and evaluated their antitumor activity in vitro and in vivo.

Results: We established an expression hierarchy for the analyzed antigens (B7-H3 = GD2 >> IL13Rα2 > HER2 = EphA2) and demonstrated that antigen expression is heterogenous. All high-grade gliomas expressed HLA class-I, but only 57.1% of other tumor subtypes had detectable expression. We then selected B7-H3 as a target for CAR T-cell therapy. B7-H3-CAR T-cells recognized tumor cells in an antigen-dependent fashion. Local or systemic administration of B7-H3-CAR T-cells induced tumor regression in PDOX and immunocompetent murine glioma models resulting in a significant survival advantage.

Conclusions: Our study highlights the importance of studying target antigen and HLA class-I expression in PDOX samples for the future design of immunotherapies. In addition, our results support active preclinical and clinical exploration of B7-H3-targeted CAR T-cell therapies for a broad spectrum of pediatric brain tumors.

Keywords: B7-H3; CAR T-cell immunotherapy; GD2; HLA; immunocompetent and PDOX models; pediatric brain tumor.

Pulmonary invasive mucinous adenocarcinoma (IMA) is considered a variant of lung adenocarcinomas based on the current World Health Organization classification of lung tumors. However, the molecular mechanism driving IMA development and progression is not well understood. Thus, we surveyed the genomic characteristics of IMA in association with immune-checkpoint expression to investigate new potential therapeutic strategies. Tumor cells were collected from surgical specimens of primary IMA, and sequenced to survey 53 genes associated with lung cancer. The mutational profiles thus obtained were compared in silico to conventional adenocarcinomas and other histologic carcinomas, thereby establishing the genomic clustering of lung cancers. Immunostaining was also performed to compare expression of programmed death ligand 1 (PD-L1) and B7-H3 in IMA and conventional adenocarcinomas. Mutations in Kirsten rat sarcoma viral oncogene homolog (KRAS) were detected in 75% of IMAs, but in only 11.6% of conventional adenocarcinomas. On the other hand, the frequency of mutations in epidermal growth factor receptor (EGFR) and tumor protein p53 (TP53) genes was 5% and 10%, respectively, in the former, but 48.8% and 34.9%, respectively, in the latter. Clustering of all 78 lung cancers indicated that IMA is distinct from conventional adenocarcinoma or squamous cell carcinoma. Strikingly, expression of PD-L1 in ≥1% of cells was observed in only 6.1% of IMAs, but in 59.7% of conventional adenocarcinomas. Finally, 42.4% and 19.4% of IMAs and conventional adenocarcinomas, respectively, tested positive for B7-H3. Although currently classified as a variant of lung adenocarcinoma, it is also reasonable to consider IMA as fundamentally distinct, based on mutation profiles and genetic clustering as well as immune-checkpoint status. The immunohistochemistry data suggest that B7-H3 may be a new and promising therapeutic target for immune checkpoint therapy.

Meningioma is the most common brain tumor in adults. Surgical resection remains the primary treatment. No chemotherapy exists. However, gene mutations now could explain ~ 80% of meningioma and targeted therapies based on these are being investigated. Furthermore, with the recent discovery of PD-L1 in malignant meningioma, clinical trials using immunotherapy have commenced. Here, we report for the first time the expression profiles of immune checkpoint proteins PD-L2, B7-H3 and CTLA-4 in meningioma and their association to common gene mutations. PD-L2 and B7-H3 expression was significantly greater than all immune checkpoint proteins studied, and particularly elevated in patients with gene mutations affecting the PI3K/AKT/mTOR pathway. CTLA-4 expressing CD3+ lymphocytes were observed in atypical and malignant meningioma and tumors harboring a PIK3CA or SMO mutation. These results identify novel targets for immunotherapy irrespective of grade and distinguish potential patient populations based on genetic classification for stratification into checkpoint inhibitor clinical trials.

Purpose: Patients with relapsed pediatric solid malignancies have few therapeutic options, and many of these patients die of their disease. B7-H3 is an immune checkpoint protein encoded by the CD276 gene that is overexpressed in many pediatric cancers. Here, we investigate the activity of the B7-H3-targeting antibody-drug conjugate (ADC) m276-SL-PBD in pediatric solid malignancy patient-derived and cell line-derived xenograft (PDX and CDX) models.

Experimental design: B7-H3 expression was quantified by RNA sequencing and by immunohistochemistry on pediatric PDX microarrays. We tested the safety and efficacy of m276-SL-PBD in two stages. Randomized trials of m276-SL-PBD of 0.5mg/kg on days 1, 8, and 15 compared to vehicle were performed in PDX or CDX models of Ewing sarcoma (N=3), rhabdomyosarcoma (N=4), Wilms tumors (N=2), osteosarcoma (N=5) and neuroblastoma (N=12). We then performed a single mouse trial (SMT) in 47 PDX or CDX models using a single 0.5 m/kg dose of m276-SL-PBD.

Results: The vast majority of PDX and CDX samples studied showed intense membranous B7-H3 expression (median H-score 177, SD 52). In the randomized trials, m276-SL-PBD showed a 92.3% response rate, with 61.5% of models showing a maintained complete response (MCR). These data were confirmed in the single mouse trial with an overall response rate of 91.5% and MCR rate of 64.4%. Treatment-related mortality rate was 5.5% with late weight loss observed in a subset of models dosed weekly x 3.

Conclusions: m276-SL-PBD has significant anti-tumor activity across a broad panel of pediatric solid tumor PDX models.

The objective includes radiofrequency ablation (RFA) of a cancerous nodule results in immunogenic cell death. Tumor antigens are presented and the inflammatory environment may help stimulate adaptive and innate antitumor immunity. The objective of this study was to investigate the immune response following RFA and subsequent surgical resection in early stage non-small cell lung cancer (NSCLC). In methods, a single-session approach of computed tomography-guided tumor biopsy with immediate frozen section (and proof of NSCLC) was performed followed by RFA of the tumor in 4 patients with a solitary pulmonary nodule. Blood samples were collected before RFA and 3 days thereafter. All patients underwent radical surgical resection by video-assisted thoracoscopic lobectomy 8 days following RFA. In results, intense infiltrations of CD4+ and CD8+ lymphocytes were found along the perimeter of the RFA-treated tumor tissue, whereas the central tumor areas remained devoid of lymphocytes. In the peripheral blood, the frequency of proinflammatory, immunostimulatory IFNγ-secreting, and immunostimulatory BDCA-3+/B7-H3- dendritic cells increased after RFA. Furthermore, a significant increase in T-cell proliferation was detected in T-cell assays after RFA and tumor resection. In this article, a local and systemic immune response subsequent to RFA and complete surgical resection in patients with NSCLC was identified for the first time. Treatment of patients with NSCLC with RFA and surgery leads to an activated and highly T-cell-stimulatory phenotype of dendritic cells, which may promote long-term immunity against NSCLC. The data suggest that the RFA-induced necrotic tumor debris can serve as an in situ antigen source to induce an autologous antitumor immune response.

B7-H3 protein is an important tumor antigen, but the expression of its isoforms, 4IgB7-H3 and 2IgB7-H3, in tumor tissues remains unknown due to the lack of specific monoclonal antibodies (mAbs). In the present study, a mAb (9C3) specifically recognizing 2IgB7-H3, but not 4IgB7-H3, was prepared. Using 9C3 and a previously prepared mAb (4H7) that recognizes 4IgB7-H3 and 2IgB7-H3, the differential expression of 2IgB7-H3 and 4IgB7-H3 was analyzed in a variety of tumor cell lines by flow cytometry. It was found that 4IgB7-H3 had a more broad spectrum of expression among the cell lines compared with 2IgB7-H3. The expression of the two isoforms was further examined in glioma tissues using reverse transcription-polymerase chain reaction and immunohistochemistry techniques. The data revealed that 2IgB7-H3, but not 4IgB7-H3, was specifically expressed in glioma. Taken together, these results demonstrated for the first time that 2IgB7-H3 is a valuable biomarker for the diagnosis of glioma.

B7-H3, a newly identified B7 family member, has functional duality as a co-stimulator and co-inhibitor that fine-tunes T cell-mediated immune responses. Given that B7-H3 expression on human monocytes and dendritic cells is enhanced by inflammatory cytokines, its potential inmmunoregulatory role at sites of inflammation has been suggested. Further, monocytes play crucial roles in the pathophysiology of various inflammatory disorders including autoimmune diseases; however, the immunological role of B7-H3 in rheumatoid arthritis (RA) has not been defined. Thus, we aimed to investigate the possible roles of monocyte B7-H3 in the pathogenesis of RA. Synovial monocytes, but not peripheral monocytes, in RA patients predominantly express surface B7-H3. The 4Ig isoform of B7-H3 is exclusively induced on the cell surface, whereas the 2Ig B7-H3 isoform is constitutively expressed in the intracytoplasmic region of both peripheral and synovial monocytes. B7-H3 knockdown experiments reveal that surface B7-H3 has an inhibitory effect on IFN-γ production in CD4 memory cells. Moreover, surface B7-H3 expression on synovial monocytes inversely correlates with RA clinical parameters. Our findings demonstrate that activation-induced B7-H3 expression on synovial monocytes has the potential to inhibit Th1-mediated immune responses and immunomodulatory roles affecting RA pathogenesis.

Colorectal cancer (CRC) is one of the most common malignancies, and chemoresistance is one of the key obstacles in the clinical outcome. Here, we studied the function of B7-H3 in regulating cell cycle-mediated chemoresistance in CRC. The ability of B7-H3 in regulating chemoresistance was investigated via cell viability, clonogenicity, apoptosis and cycle analysis in vitro. Moreover, the role of B7-H3/CDC25A axis in regulating chemoresistance in vivo in the xenograft tumor models by intraperitoneal injection of oxaliplatin (L-OHP) and CDC25A inhibitors. The correlation between B7-H3 and CDC25A was examined in the CRC patients by immunohistochemistry (IHC) and pathological analyses. We found that B7-H3 could effectively enhance the resistance to a chemotherapeutic drug (oxaliplatin or 5-fluorouracil) via CDC25A. B7-H3 regulated the expression of CDC25A by the STAT3 signaling pathway in CRC cells. Furthermore, overexpression of B7-H3 enhanced chemoresistance by reducing the G2/M phase arrest in a CDC25A-dependent manner. Silencing CDC25A or treatment with CDC25A inhibitor could reverse the B7-H3-induced chemoresistance of cancer cells. Moreover, both B7-H3 and CDC25A were significantly upregulated in CRC samples compared with normal adjacent tissues and that the levels correlated with tumor stage. CDC25A was positively correlated with B7-H3 expression in this cohort. Taken together, our findings provide an alternative mechanism by which CRC cells can acquire chemoresistance via the B7-H3/CDC25A axis.

Extranodal nasal natural killer (NK)/T cell lymphoma (ENKTCL) is a rare but highly aggressive subtype of non-Hodgkin lymphoma (NHL). Nevertheless, despite extensive research, the estimated 5-year overall survival of affected patients remains low. Therefore, new treatment strategies are needed urgently. Recent advances in immunotherapy have the potential to broaden the applications of chimeric antigen receptor-modified T (CAR-T) cells and the bispecific T-cell engaging (BiTE) antibody. Here, we screened a panel of biomarkers including the B7-H3, CD70, TIM-3, VISTA, ICAM-1, and PD-1 in NKTCL cell lines. As a result, we found for the first time that B7-H3 was highly and homogeneously expressed in these cells. Consequently, we constructed a novel anti-B7-H3/CD3 BiTE antibody and B7-H3-redirected CAR-T cells, and evaluated their efficacy against NKTCL cel lines both in vitro and in vivo. Notably, we found that both anti-B7-H3/CD3 BiTE and B7-H3-redirected CAR-T cells effectively targeted and killed NKTCL cells in vitro, and suppressed the growth of NKTCL tumors in NSG mouse models. Thus, B7-H3 might be a promising therapeutic target for treating patients with NKTCL tumors.

Immunosurveillance is a critical mechanism guarding against tumor development and progression. Checkpoint inhibitors have shown significant success in cancer treatment, but expression of key factors such as PD-L1 in putative cancer stem cell (CSC) populations in squamous cell carcinoma has been inconclusive, suggesting that CSCs may have developed other mechanisms to escape immune surveillance. Here we show that CSCs upregulate the immune checkpoint molecule CD276 (B7-H3) to evade host immune responses. CD276 is highly expressed by CSCs in mouse and human head and neck squamous cell carcinoma (HNSCC) and can be used to prospectively isolate tumorigenic CSCs. Anti-CD276 antibodies eliminate CSCs in a CD8⁺ T cell-dependent manner, inhibiting tumor growth and lymph node metastases in a mouse HNSCC model. Single-cell RNA sequencing (RNA-seq) showed that CD276 blockade remodels SCC heterogeneity and reduces epithelial-mesenchymal transition. These results show that CSCs utilize CD276 for immune escape and suggest that targeting CD276 may reduce CSCs in HNSCC.

Keywords: BMI1; CD276; cancer stem cell; head and neck squamous cell carcinoma; immune surveillance; lineage tracing.

Tumor angiogenesis is required for tumor growth and metastasis, and the Ang/Tie-2 axis plays a pivotal role in angiogenesis. B7-H3, a new member of the B7 family of costimulatory molecules, has a critical function in the T-cell-mediated antitumor immune response, and abnormal tumor B7-H3 expression is frequently associated with a poor prognosis. However, the relationship between B7-H3 and angiogenesis in clear cell renal carcinoma (ccRCC) remains unclear. In this study, we used immunohistochemical methods to detect tumor vascular expression of B7-H3 and Tie-2 in tissue microarrays of 82 ccRCC patient samples. According to the results, B7-H3 is highly expressed in the tumor vascular endothelium of ccRCC and is associated with the ccRCC grade and tumor-node-metastasis (TNM) stage. Although vascular Tie-2 expression was also correlated with T stage and lymph node metastasis, it was not related to ccRCC grade or distant metastasis. The microvessel density (MVD) labeled by CD34 was correlated with tumor grade and TNM stage. Expression of B7-H3 and Tie-2 was positively correlated, and the levels were positively associated with the MVD. Additionally, immunofluorescence staining revealed coexpression of B7-H3 and Tie-2 in the vascular endothelia of ccRCC. Collectively, our findings suggest that expression of B7-H3 and Tie-2 in ccRCC tumor vasculature is closely related to the progression and prognosis of the disease. Furthermore, B7-H3 possibly promotes ccRCC angiogenesis through the Tie-2 pathway. Thus, B7-H3 might serve as an effective endothelial marker for ccRCC prognosis and become a promising target for ccRCC anti-angiogenic-targeted therapy.

To characterize the expression of co-signaling molecules PD-L1, PD-1, and B7-H3 in cutaneous squamous cell carcinoma (cSCC) by immune status.

We retrospectively analyzed 66 cases of cSCC treated with surgical resection from 2012 to 2015. Immunostained tumor sections were analyzed for percent of tumor cells expressing PD-L1 (Tum-PD-L1%), B7-H3 (Tum-B7-H3%), density of peri and intratumoral CD8 T cells (CD8 density), proportion of CD8 T cells expressing PD-1 (CD8-PD-1%) and of tumor-infiltrating immune cells (TII) expressing PD-L1 (TII-PD-L1%).

Of 66 cases, 42 were immunocompetent, 24 immunosuppressed (13 organ transplant, 8 HIV+, 3 other). Defining positive expression at>> 5%, 26% of tumors were positive for PD-L1, 85% for B7-H3, 80% had CD8 T cells that expressed PD-1 and 55% had TII that expressed PD-L1. Tum-B7-H3% was significantly higher (median 60 vs. 28%, p = 0.025) in immunocompetent vs. immunosuppressed patients, including when factoring in cause of immunosuppression. No significant difference in Tum-PD-L1%, TII-PD-L1%, CD8 density, or CD8-PD-1% was observed. Tumors from HIV+ patients lacked PD-L1 expression, and had lower B7-H3% (median 2.5 vs. 60%, p = 0.007), and higher CD8 density (median 75% vs. 40%, p = 0.04) compared to immunocompetent patients. Higher tumor grade (Rs = 0.34, p = 0.006) and LVI (Rs = 0.61, p < 0.001) were both associated with higher Tum-PD-L1%.

cSCC showed expression of PD-L1 on tumor in 26% of cases, and high tumor B7-H3 expression (85%) and PD-1 expression on CD8 TILs (80%). Tumor B7-H3 expression was significantly higher in immunocompetent vs. immunosuppressed patients, largely driven by very low expression in HIV+ patients.

Over the past few decades, different inhibitory receptors have been identified, which have played prominent roles in reducing anti-tumor immune responses. The role of immune checkpoint inhibitors in cancer was revealed by critical blockade of the cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) and programmed cell death protein-1 (PD-1) checkpoints. Immune checkpoint inhibitors, including anti-PD-1 (nivolumab and pembrolizumab), anti-PD-L1 (Atezolizumab, avelumab, and duravulumab), and anti-CTLA-4 (ipilimumab, tremelimumab), are currently FDA-approved treatment options for a broad range of cancer types. However, regarding immunotherapy advances in recent years, most studies have been focused on finding the antibodies against other inhibitory immune checkpoints in the tumor microenvironment such as lymphocyte activation gene-3 (LAG-3), T cell immunoglobulin, and mucin domain 3 (TIM-3), B7-homolog 3 (B7-H3), V-domain immunoglobulin-containing suppressor of T-cell activation (VISTA), diacylglycerol kinase-α (DGK-α), T cell immunoglobulin and ITIM domain (TIGIT), and B and T lymphocyte attenuator (BTLA). This immune checkpoint exerts differential inhibitory impacts on various types of lymphocytes. The suppression of immune responses demonstrates a surprising synergy with PD-1. Therefore, most antibodies against these immune checkpoints are undertaking clinical trials for cancer immunotherapy of advanced solid tumors and hematologic malignancies. In this review, we will summarize recent findings of immune checkpoint and the role of monoclonal antibodies in cancer immunotherapy targeting these receptors.

Keywords: Cancer; Immune checkpoint; Immunotherapy; Inhibitory receptor.