Immunoregulatory protein B7-H3 upregulated in bacterial and viral infection and its diagnostic potential in clinical settings.

Bacterial and viral infections cause a huge burden to healthcare settings worldwide, and mortality rates associated with infectious microorganisms have remained high in recent decades. Despite tremendous efforts and resources worldwide to explore diagnostic biomarkers, rapid and easily assayed indicators for the diagnosis of bacterial and viral infections remain a challenge. B7 homolog 3 (B7-H3), a member of the B7 family of immunoregulatory proteins, is overexpressed in patients with septicemia, meningitis, pneumonia, and hepatitis. Therefore, B7-H3 could be used as a potential clinical indicator and therapeutic target for bacterial and viral infections caused by H. pylori, S. pneumoniae, M. pneumoniae, hepatitis B virus (HBV), viral hemorrhagic septicemia virus (VHSV), respiratory syncytial virus (RSV), and human immunodeficiency virus (HIV). Moreover, the interplay between infectious microorganisms and B7-H3 and exploration of the functional roles of the B7-H3 molecule could aid in the development of novel strategies for disease diagnosis and immunotherapy.

Ultrasound Molecular Imaging Enhances High-Intensity Focused Ultrasound Ablation on Liver Cancer With B7-H3-Targeted Microbubbles.

BACKGROUND: High-intensity focused ultrasound (HIFU) is a promising minimally invasive treatment for liver cancer; however, its efficacy is often limited by the attenuation of ultrasonic energy. This study investigates the effectiveness of B7-H3-targeted microbubbles (T-MBs) in enhancing HIFU ablation of liver cancer and explores their potential for clinical translation. METHODS: T-MBs and isotype control microbubbles (I-MBs) were synthesized through the conjugation of biotinylated anti-B7-H3 antibody and isotype control antibody to the microbubble surface, respectively. Contrast-enhanced ultrasound imaging was performed to compare the accumulation of T-MBs and I-MBs in liver cancer at various time points. The efficacy of T-MBs in enhancing HIFU treatment was evaluated by measuring the immediate tumor ablation rate and long-term tumor growth suppression. Additionally, the induced antitumor immune response was assessed through cytokine quantification in serum and tumor tissue, along with immunofluorescence staining conducted on days 1, 3, and 7 post-treatment. RESULTS: T-MBs demonstrated superior liver cancer-specific accumulation, characterized by higher concentrations and prolonged retention compared to I-MBs. The combination of T-MBs with HIFU resulted in significantly enhanced tumor ablation rates and superior tumor growth suppression. Post-treatment analysis revealed a gradual uptick in cytokine levels within the tumor microenvironment, along with progressive infiltration of antitumor immune cells. CONCLUSION: T-MBs effectively enhance the therapeutic efficacy of HIFU for liver cancer treatment while simultaneously promoting an antitumor immune response. These findings provide a strong experimental foundation for the clinical translation of ultrasound molecular imaging combined with HIFU as a novel approach for tumor therapy.

Targeting CD276 for T cell-based immunotherapy of breast cancer.

BACKGROUND: Breast cancer (BC) is the most common malignancy in women. Immunotherapy has revolutionized treatment options in many malignancies, and the introduction of immune checkpoint inhibition yielded beneficial results also in BC. However, many BC patients are ineligible for this T cell-based therapy, others do not respond or only briefly. Thus, there remains a high medical need for new therapies, particularly for triple-negative BC. CD276 (B7-H3) is overexpressed in several tumors on both tumor cells and tumor vessels, constituting a promising target for immunotherapy. METHODS: We analyzed tumor samples of 25 patients using immunohistochemistry to assess CD276 levels. The potential of CC-3, a novel bispecific CD276xCD3 antibody, for BC treatment was evaluated using various functional in vitro assays. RESULTS: Pronounced expression of CD276 was observed in all analyzed tumor samples including triple negative BC. In analyses with BC cells, CC-3 induced profound T cell activation, proliferation, and T cell memory subset formation. Moreover, treatment with CC-3 induced cytokine secretion and potent tumor cell lysis. CONCLUSION: Our findings characterize CD276 as promising target and preclinically document the therapeutic potential of CC-3 for BC treatment, providing a strong rationale for evaluation of CC-3 in BC patients in a clinical trial for which the recruitment has recently started.

Antitumor effects of intracranial injection of B7-H3-targeted Car-T and Car-Nk cells in a patient-derived glioblastoma xenograft model.

BACKGROUND: Glioblastoma multiforme (GBM) is the most lethal primary brain tumor for which novel therapies are needed. Recently, chimeric antigen receptor (CAR) T cell therapy has been shown to be effective against GBM, but it is a personalized medicine and requires high cost and long time for the cell production. CAR-transduced natural killer (NK) cells can be used for "off-the-shelf" cellular immunotherapy because they do not induce graft-versus-host disease. Therefore, we aimed to analyze the anti-GBM effect of CAR-T or NK cells targeting B7-H3, which is known to be highly expressed in GBM. METHODS: CAR-T cells targeting B7-H3 were generated using previously reported anti-B7-H3 scFv sequences. Cord blood (CB)-derived NK cells transduced with the B7-H3 CAR were also generated. Their anti-GBM effect was analyzed in vitro. The antitumor effect of intracranial injection of the B7-H3 CAR-T or NK cells was investigated in an in vivo xenograft model with patient-derived GBM cells. RESULTS: Both B7-H3 CAR-T cells and CAR-NK cells exhibited marked cytotoxicity against patient-derived GBM cells in vitro. Furthermore, intracranial injection of CAR-T cells and CAR-NK cells targeting B7-H3 resulted in a significant antitumor effect against patient-derived GBM xenografts. CONCLUSION: Not only CAR-T cells but also CB-derived CAR-NK cells targeting B7-H3 may have the potential to eliminate GBM cells.

B7-H3 is widely expressed in soft tissue sarcomas.

PURPOSE: Targeted therapy development in soft tissue sarcoma (STS) has been burdened by the heterogeneity of this group of rare tumors. B7 homolog 3 protein (B7-H3) is a molecule in the same family as programmed death-ligand 1 (PD-L1). It has limited expression in noncancerous tissues and is overexpressed in many cancers, making it an attractive target for cancer therapy, and clinical trials targeting B7-H3 are actively underway. While available data demonstrate high expression levels of B7-H3 in individual sarcoma subtypes, its expression patterns across STS subtypes are not well described. The purpose of this study was to characterize the expression patterns of B7-H3 in STS. PATIENTS AND METHODS: This retrospective analysis evaluated STS tumor specimens from patients with a variety of different subtypes. Specimens were evaluated by immunohistochemistry (IHC) for expression and staining pattern of B7-H3 both in tumors and in associated vasculature. RESULTS: Specimens from 153 sarcoma patients included 15 different STS subtypes. B7-H3 was broadly expressed in 97% of samples (95% CI 0.93-0.99) and 69.2% demonstrated high levels of B7-H3 expression (95% CI 0.61-0.76). No significant association between B7-H3 positivity or expression level and prior treatment(s), tumor size, tumor grade, or patient age. B7-H3 positivity in vessels was found in 94.7% (145/153) of samples. In tumors that had been previously assessed for PD-L1 and PD-1, there was no correlation between B7-H3 positivity or expression and the positivity or expression level of PD-L1 or PD-1. CONCLUSION: These data show high levels of B7-H3 positivity across soft tissue sarcoma subtypes, suggesting its feasibility as a therapeutic target for future sarcoma treatments. Future clinical trials are needed to evaluate whether targeting B7-H3 can provide clinical benefit to help patients with sarcoma.

B7-H3 promotes nasopharyngeal carcinoma progression by regulating CD8+ T cell exhaustion.

BACKGROUND: B7-H3 protein is an important regulator of the adaptive immune response in human tumorigenesis. 4-1BB is a co-stimulatory receptor expressed on activated CD8+ T cells, and regulates T cell immunity. Here, we investigated the role of B7-H3 in the growth and invasion of nasopharyngeal carcinoma (NPC) and the effect of its interaction with 4-1BB on tumor immunity. METHODS: Short hairpin (sh) RNA was designed to knock down B7-H3 expression in NPC cells. NPC cells with stable knockdown of B7-H3 were established and injected into nude mice. The effects of B7-H3 on cell proliferation, apoptosis, and epithelial-to-mesenchymal transition (EMT) were detected by the CCK8 assay, flow cytometry, TUNEL assay, and western blot analysis. The migration and invasion abilities were determined using the Transwell assay and scratch assay. Co-immunoprecipitation (Co-IP) assays were performed to study the interaction between B7-H3 and 4-1BB. Anti-4-1BB antibody was used in a co-culture system and xenograft mice to study the effect of 4-1BB on NPC development. RESULTS: NPC cells transfected with sh-B7-H3 showed a higher rate of apoptosis, slower growth rate, impaired migration, and less EMT in vitro. Xenograft mice with stable knockout of B7-H3 had lower tumor burdens, and the stripped tumors had lower rates of cell proliferation, higher rates of apoptosis, and less EMT in vivo. Additionally, decreased B7-H3 expression was positively correlated with interferon-γ, tumor necrosis factor-α, and 4-1BB+CD8+ tumor-infiltrating lymphocytes. Co-IP studies showed that B7-H3 interacts with 4-1BB. Also, the inhibitory effects of sh-B7-H3 on NPC tumor growth, invasion, and tumor immunity could be alleviated by the anti-4-1BB antibody both in vivo and in vitro. CONCLUSION: Our findings suggest that B7-H3 may accelerate tumor growth, tumor cell invasion, and EMT, and interact with 4-1BB to produce CD8+ T cell exhaustion that inhibits tumor immunity. B7-H3 might serve as a novel target for treating NPC.

B7H3 Immune Checkpoint Overexpression Is Associated with Decreased Complete Response Rates to Neoadjuvant Therapy in Locally Advanced Rectal Cancer.

BACKGROUND AND OBJECTIVES: Rectal cancer accounts for approximately one-third of colorectal cancers, with over 340,000 deaths globally in 2022. Despite advancements in treatment, the five-year overall survival for locally advanced rectal cancer (LARC) remains at 74%, with significant morbidity. B7H3 (CD276), an immune checkpoint protein, plays a role in tumor progression and resistance to therapy, and correlates with poor prognosis in various cancers, including colorectal cancer. This study aims to evaluate the expression of B7H3 in LARC and its impact on overall complete response (oCR) rates to neoadjuvant therapy. METHODS: A retrospective study was conducted on 60 patients with LARC who received neoadjuvant chemoradiation (nCRT) followed by total mesorectal excision (TME). B7H3 expression was assessed using immunohistochemistry on surgical specimens. Expression levels were categorized as high or low based on a composite score, and their association with oCR rates was analyzed. RESULTS: High B7H3 expression was observed in 60% of patients, with 73.5% showing expression in more than 50% of tumor cells. Patients who achieved oCR had significantly lower B7H3 expression compared to those with residual disease (p < 0.001). No nuclear expression of B7H3 was detected. No significant correlation was found between B7H3 expression and other clinicopathological variables, except for a higher likelihood of non-restorative surgery in patients with elevated B7H3 levels (p = 0.049). Mucinous adenocarcinoma had high expression of B7H3. CONCLUSIONS: Elevated B7H3 expression is associated with reduced oCR rates in LARC, highlighting its potential role as a prognostic biomarker. Further studies with larger cohorts are warranted to validate these findings and explore B7H3-targeted therapies as a treatment strategy for LARC.

Development of a Specifically Labeled 89Zr Antibody for the Noninvasive Imaging of Tumors Overexpressing B7-H3.

B7-H3 has emerged as a promising target and potential biomarker for diagnosing tumors, evaluating treatment efficacy, and determining patient prognosis. Hu4G4 is a recombinant humanized antibody that selectively targets the extracellular domain of human B7-H3. In this study, we describe the radiolabeling of hu4G4 with the positron emission tomography (PET) emitter radionuclide zirconium 89 (89Zr) and evaluate its potency as an immuno-PET tracer for B7-H3-targeted imaging by comparing it in vitro and in vivo to [89Zr]Zr-DFO-DS-5573a using various models. The radiolabeled compound, [89Zr]Zr-desferrioxamine-hu4G4 ([89Zr]Zr-DFO-hu4G4), demonstrated a high radiochemical purity (RCP) of greater than 99% and a specific activity of 74 MBq/mg following purification. Additionally, it maintained stability in human serum albumin (HSA) and acetate buffer, preserving over 90% of its RCP after 7 days. Three cell lines targeting human B7-H3(U87/CT26-CD276/GL261-CD276) were used. Flow cytometry analysis indicated that the B7-H3-positive cells (U87/CT26-CD276/GL261-CD276) had a higher B7-H3 protein level with no expression in the B7-H3-negative cells (CT26-wt/GL261-wt) (P < 0.001). Moreover, the cellular uptake was 45.71 ± 3.78% for [89Zr]Zr-DFO-hu4G4 in CT26-CD276 cells versus only 0.93 ± 0.47% in CT26-wt cells and 30.26 ± 0.70% when [89Zr]Zr-DFO-hu4G4 in CT26-CD276 cells were blocked with 100× 8H9. The cellular uptake of [89Zr]Zr-DFO-hu4G4 was akin to that observed with [89Zr]Zr-DFO-DS-5573a with no significant differences (45.71 ± 3.78 % vs 47.07 ± 0.86 %) in CT26-CD276 cells. Similarly, the CT26-CD276 mouse model demonstrated markedly low organ uptake and elevated tumor uptake 48 h after [89Zr]Zr-DFO-hu4G4 injection. PET/CT analysis showed that the tumor-to-muscle (T/M) ratios were substantially higher compared to other imaging groups: 27.65 ± 3.17 in CT26-CD276 mice versus 11.68 ± 4.19 in CT26-wt mice (P < 0.001) and 16.40 ± 0.78 when 100× 8H9 was used to block [89Zr]Zr-DFO-hu4G4 in CT26-CD276 mice (P < 0.01) at 48 h post-injection. Additionally, the tracer showed markedly high accumulation in the tumor region (22.57 ± 3.03% ID/g), comparable to the uptake of [89Zr]Zr-DFO-DS-5573a (24.76 ± 5.36% ID/g). A dosimetry estimation study revealed that the effective dose for [89Zr]Zr-DFO-hu4G4 was 2.96 × 10-01 mSv/MBq, which falls within the acceptable range for further research in nuclear medicine. Collectively, these results indicated that [89Zr]Zr-DFO-hu4G4 was successfully fabricated and applied in B7-H3-targeted tumor PET/CT imaging, which showed excellent imaging quality and tumor detection efficacy in tumor-bearing mice. It is a promising imaging agent for identifying tumors that overexpress B7-H3 for future clinical applications.

Therapeutic vaccine targeting dual immune checkpoints induces potent multifunctional CD8+ T cell anti-tumor immunity.

BACKGROUND: Vaccines targeting immune checkpoints represent a promising immunotherapeutic approach for solid tumors. However, the therapeutic efficacy of dual targeting immune checkpoints is still unclear in renal carcinoma. METHODS: An adenovirus (Ad) vaccine targeting B7H1 and B7H3 was developed and evaluated for its therapeutic efficacy in subcutaneous, lung metastasis or orthotopic renal carcinoma mouse and humanized models using flow cytometry, Enzyme-linked immunosorbent spot (ELISPOT), cytotoxic T lymphocyte (CTL) killing, cell deletion, hematoxylin and eosin (HE) staining, and immunohistochemistry (IHC) assays. RESULTS: The Ad-B7H1/B7H3 immunization effectively inhibited tumor growth and increased the induction and percentages of CD8+ T cells in subcutaneous tumor models. The vaccine enhanced the induction and maturation of CD11c+ or CD8+CD11c+ cells, promoting tumor-specific CD8+ T cell immune responses. This was evidenced by increased proliferation of CD8+ T cells and enhanced CTL killing activity. Deletion of CD8+ T cells in vivo abolished the anti-tumor effect of the Ad-B7H1/B7H3 vaccine, highlighting the pivotal role of functional CD8+ T cell immune responses. Moreover, significant therapeutic efficacy of the Ad-B7H1/B7H3 vaccine was observed in lung metastasis, orthotopic, and humanized tumor models through multifunctional CD8+ T cell immune responses. CONCLUSIONS: The Ad vaccine targeting dual immune checkpoints B7H1 and B7H3 exerts a potent therapeutic effect for renal carcinoma and holds promise for solid tumor treatment.

Multiplexed Spatial Imaging at the Single-Cell Level Reveals Mutually Exclusive Expression of B7 Family Proteins.

Targeting novel inhibitory ligands beyond anti-PD-1 and PD-L1 and CTLA-4 therapies is essential for the next decade of the immunotherapy era. Agents for the B7 family molecules B7-H3, B7-H4, and B7-H5 are emerging in clinical trial phases; therefore, further accumulation of evidence from both clinical and basic aspects is vital. Here, we applied a 7-color multiplexed imaging technique to analyze the profile of B7 family B7-H3/B7-H4/B7-H5 expression, in addition to PD-L1, and the spatial characteristics of immune cell infiltrates in urothelial carcinoma (UC). The results revealed that B7-H3 and B7-H4 were mainly expressed on tumor cells and B7-H5 on immune cells in UC, and most of the B7-H3/B7-H4/B7-H5-positive cells were mutually exclusive with PD-L1-positive cells. Also, the expression of B7-H4 was elevated in patients with advanced pathologic stages, and high B7-H4 expression was a significant factor affecting overall mortality following surgery in UC. Furthermore, spatial analysis revealed that the distance from the B7-H4+ cells to the nearest CD8+ cells was markedly far compared with other B7 family-positive tumor cells. Interestingly, the distance from B7-H4+ cells to the nearest CD8+ cells was significantly farther in patients dying from cancer after surgery or immune checkpoint inhibitors compared with cancer survivors; thus, high B7-H4 expression in tumor cells may inhibit CD8 infiltration into the tumor space and that B7-H4-positive cells form a specific spatial niche. In summary, we performed a comprehensive evaluation of B7 family member expression and found that the spatial distribution of B7-H4 suggests the potentially useful role of combination blockade with both B7-H4 and the current anti-PD-1/PD-L1 axis in the treatment of UC.

PD-L1 and B7-H3 are Effective Prognostic Factors and Potential Therapeutic Targets for High-Risk Thyroid Cancer.

The prognosis of thyroid cancer in patients varies significantly based on different pathological types or distinct clinical situations. Investigating the expression of immune checkpoint molecules PD-L1 and B7-H3 in high-risk thyroid cancer and their correlation with clinicopathological features and prognosis will contribute to the development of novel therapeutic strategies. A retrospective sample of 202 patients with thyroid cancer who underwent surgery at the Cancer Hospital of the Chinese Academy of Medical Sciences was collected, including 33 cases of anaplastic thyroid cancer (ATC), 21 cases of differentiated thyroid cancer (DTC) with distant metastasis (DM), 7 cases of differentiated high-grade thyroid carcinoma (DHGTC), and 109 cases of aggressive subtypes of papillary thyroid carcinoma (PTC) (including 28 cases of tall cell PTC, 31 cases of diffuse sclerosing PTC, 20 cases of solid PTC, 15 cases of columnar cell PTC, and 15 cases of hobnail PTC). In the control group, there were 32 cases of classic PTC. The differences in protein expression between PD-L1 and B7-H3 in several high-risk thyroid cancers and normal tissues and controls were compared by immunohistochemical staining, and the clinicopathological features and prognostic relevance were statistically analyzed. The expression of PD-L1 in ATC (P < 0.001), tall cell PTC (P = 0.031), and DHGTC (P = 0.003) was significantly higher than that in classic PTC. The expression of B7-H3 in ATC (P < 0.001), DTC with DM (P = 0.001), diffuse sclerosing PTC (P = 0.013), columnar cell PTC (P = 0.007), solid PTC (P < 0.001), hobnail PTC (P < 0.001), and DHGTC (P < 0.001) was significantly higher than that in classic PTC. In ATC, PD-L1 expression correlated significantly with extrathyroidal extension (ETE) (P = 0.027) and B7-H3 expression correlated significantly with male patients (P = 0.031) and lymph node metastasis (LNM) (P = 0.026). The positive expression of B7-H3 (P = 0.041) was an independent risk factor for disease progression in ATC. B7-H3 positive expression (P = 0.049), PD-L1 positive expression (P = 0.015), and tumor diameter ≥ 2 cm (P = 0.038) were independent risk factors for disease progression in patients with DTC with DM. PD-L1 positive expression (P = 0.019) and tumor diameter ≥ 2 cm (P = 0.018) were independent risk factors for disease progression in patients with aggressive subtypes of PTC. B7-H3 and PD-L1 are expected to be effective prognostic indicators for patients with aggressive thyroid cancer, which can help in optimization of individualized treatment strategies. Immunotherapy targeting these two molecules may provide new and complementary ideas for the treatment of high-risk/refractory thyroid cancer.

Interleukin-7 expression by CAR-T cells improves CAR-T cell survival and efficacy in chordoma.

Chordoma is a rare bone tumor that frequently recurs after surgery, and the prognosis is poor with current treatments. This study aimed to identify potential novel immunotherapeutic targets for chordomas by identifying target proteins in clinical samples as well as tumor microenvironmental factors to enhance efficacy. Fourteen chordoma samples were analyzed by single-cell RNA sequencing, and B7-H3 and IL-7 were identified as potential targets and potentiators, respectively. B7-H3-targeted chimeric antigen receptor T (CAR-T) cells and B7-H3 CAR-T cells expressing IL-7 were synthesized and their anti-tumor activity evaluated in vitro, including in primary chordoma organoid models. The B7-H3 CAR-T/IL-7 therapy showed enhanced cytotoxicity and prolonged duration of action against tumor cells. Additionally, IL-7 modulated favorable subpopulations of cultured CAR-T cells, diminished immune checkpoint expression on T-cell surfaces, and enhanced T-cell functionality. The incorporation of IL-7 molecules into the B7-H3 CAR structure augmented CAR-T-cell function and improved CAR-T-cell efficacy, thus providing a novel dual therapeutic strategy for chordoma treatment.

B7-H3 promotes the migration and invasion of colorectal cancer cells via regulating the actin cytoskeleton and RhoA/ROCK1/LIMK1 signaling pathway.

BACKGROUND AND AIMS: Aberrant expression of B7 homolog 3 protein (B7-H3) has been detected in various cancers including colorectal cancer (CRC) and implicated in modulating multiple biological functions of CRC cells. However, its role in CRC metastasis has not yet been determined. This study aims to explore and unravel the underlying mechanisms through which B7-H3 contributes to migration, invasion and actin cytoskeleton in CRC. METHODS: The expression of B7-H3 and LIMK1 in CRC tumor samples was determined by IHC staining. Transwell and F-actin immunofluorescence staining assays were performed to explore the role of B7-H3 in migration, invasion and actin filament accumulating of CRC cells. RNA-seq and Western blot assays were used to investigate the molecular mechanisms. RESULTS: B7-H3 was highly expressed in CRC tissues and positively associated with poor prognosis of CRC patients by immunohistochemistry. Migration and invasion assays showed that B7-H3 knockdown significantly inhibited the migration and invasion of CRC cells. B7-H3 overexpression had the opposite effect. Moreover, we determined that B7-H3 could regulate actin cytoskeleton and the RhoA/ROCK1/LIMK1 pathway by F-actin immunofluorescence staining and Western blot. Importantly, the BDP5290, an inhibitor of the RhoA/ROCK1/(LIM domain kinase 1) LIMK1 axis, reversed the effects of B7-H3 overexpression on actin filament accumulating, migration, and invasion of CRC cells. CONCLUSIONS: Our study concluded that B7-H3 facilitated CRC cell actin filament accumulating, migration, and invasion through the RhoA/ROCK1/LIMK1 axis.

Diet restriction enhances the effect of immune checkpoint block by inhibiting the intratumoral mTORC1/B7-H3 axis.

Immune checkpoint blockade therapy has demonstrated significant therapeutic efficacy in certain cancer types; however, the impact of dietary restriction remains scarcely reported in this context. This study aimed to investigate the influence of dietary restriction on anti-PDL-1 therapy and the interplay of immune cells within this context. Using an anti-PDL-1 regimen combined with dietary restrictions, tumor progression was assessed in LLC-bearing mice. Flow cytometry was employed to analyze immune cell infiltration and differentiation levels within the tumor microenvironment. The expression of mTORC1/B7-H3 in tumors subjected to dietary restriction was also examined. LLC tumors with elevated B7-H3 expression were validated in mice to determine its inhibitory effect on immune cell proliferation and differentiation. A CD3/B7-H3 chimeric antibody was developed for therapeutic intervention in B7-H3 overexpressing tumors, with subsequent T cell responses assessed through flow cytometry. Dietary restriction potentiated the effect of anti-PDL1 therapy by suppressing the intratumorally mTORC1/B7-H3 axis. In vivo experiments demonstrated that elevated B7-H3 expression in tumors reduced infiltration and activation of CD8 + T cells within the tumor, while it did not affect tumor-infiltrating Tregs. In vitro studies revealed that high B7-H3 expression influenced the proliferation and activation of CD8 + T cells within a Coculture system. The constructed CD3/B7-H3 chimeric antibody prominently activated TCR within B7-H3 overexpressing tumors and impeded tumor progression. The findings suggest that dietary restriction enhances the efficacy of immune checkpoint blockade by modulating the intratumoral mTORC1/B7-H3 axis.

CAR T cells redirected to B7-H3 for pediatric solid tumors: Current status and future perspectives.

Despite intensive therapies, pediatric patients with relapsed or refractory solid tumors have poor outcomes and need novel treatments. Immune therapies offer an alternative to conventional treatment options but require the identification of differentially expressed antigens to direct antitumor activity to sites of disease. B7-H3 (CD276) is an immune regulatory protein that is expressed in a range of malignancies and has limited expression in normal tissues. B7-H3 is highly expressed in pediatric solid tumors including osteosarcoma, rhabdomyosarcoma, Ewing sarcoma, Wilms tumor, neuroblastoma, and many rare tumors. In this article we review B7-H3-targeted chimeric antigen receptor (B7-H3-CAR) T cell therapies for pediatric solid tumors, reporting preclinical development strategies and outlining the landscape of active pediatric clinical trials. We identify challenges to the success of CAR T cell therapy for solid tumors including localizing to and penetrating solid tumor sites, evading the hostile tumor microenvironment, supporting T cell expansion and persistence, and avoiding intrinsic tumor resistance. We highlight strategies to overcome these challenges and enhance the effect of B7-H3-CAR T cells, including advanced CAR T cell design and incorporation of combination therapies.

Generation of B7-H3 isoform regulated by ANXA2/NSUN2/YBX1 axis in human glioma.

In recent years, in the development of emerging immunotherapy, B7-H3 is also termed as CD276 and has become a novel chimeric antigen receptor (CAR)-T target against glioma and other tumours, and aroused extensive attention. However, B7-H3 has three isoforms (2, 3 and 4Ig) with the controversial expression and elusive function in tumour especially glioma. The current study mainly focuses on the regulatory factors and related mechanisms of generation of different B7-H3 isoforms. First, we have determined that 2Ig is dominant in glioma with high malignancy, and 4Ig is widely expressed, whereas 3Ig shows negative expression in all glioma. Next, we have further found that RNA binding protein annexin A2 (ANXA2) is essential for B7-H3 isoform maintenance, but fail to determine the choice of 4Ig or 2Ig. RNA methyltransferase NOP2/Sun RNA methyltransferase 2 (NSUN2) and 5-methylcytosine reader Y-box binding protein 1 (YBX1) facilitate the production of 2Ig. Our findings have uncovered a series of factors (ANXA2/NSUN2/YBX1) that can determine the alternative generation of different isoforms of B7-H3 in glioma. Our result aims to help peers gain a clearer understanding of the expression and regulatory mechanisms of B7H3 in tumour patients, and to provide better strategies for designing B7H3 as a target in immunotherapy.

Non-immune functions of B7-H3: bridging tumor cells and the tumor vasculature.

B7-H3 (CD276), an immune checkpoint molecule, is overexpressed in various types of cancer and their tumor vasculature, demonstrating significant associations with adverse clinical outcomes. In addition to its well-known immune functions, B7-H3 exhibits dual co-stimulatory/co-inhibitory roles in normal physiology and the tumor microenvironment. The non-immune functions of B7-H3 in tumor cells and the tumor vasculature, including promoting tumor cell anti-apoptosis, proliferation, invasion, migration, drug resistance, radioresistance, as well as affecting cellular metabolism and angiogenesis, have increasingly gained attention from researchers. Particularly, the co-expression of B7-H3 in both tumor cells and tumor endothelial cells highlights the higher potential and clinical utility of therapeutic strategies targeting B7-H3. This review aims to summarize the recent advances in understanding the non-immune functions of B7-H3 in tumors and provide insights into therapeutic approaches targeting B7-H3, focusing on its co-expression in tumor cells and endothelial cells. The aim is to establish a theoretical foundation and practical reference for the development and optimization of B7-H3-targeted therapies.

Expression and Prognostic Value of a Novel B7-H3 (CD276) Antibody in Acute Myeloid Leukemia.

Despite recent advances in immunophenotyping, the prognosis of acute myeloid leukemia (AML) is still mainly estimated using age and genetic markers. As the genetic heterogeneity of AML patients is high, flow cytometry-based classification with appropriate biomarkers can efficiently complement risk stratification and treatment selection. An increased expression of B7-H3 (CD276), an immune checkpoint protein, has been reported and associated with poor prognosis. However, the available data are limited and heterogeneous. Here, we used a novel, proprietary murine anti-B7-H3 8H8 antibody for the flow cytometric analysis of B7-H3 expression in AML blasts from 77 patients. Our antibody reliably detected substantial B7-H3 expression in 62.3% of AML patients. B7-H3 expression was higher in the monocytic French-American-British (FAB) M5 group and in intermediate and poor risk patients according to the European Leukemia Network. Using receiver operating characteristics (ROCs), we identified a specific fluorescence intensity cut-off of 4.45 to discriminate between B7-H3high and B7-H3low expression. High B7-H3 expression was associated with shorter overall survival (OS) and progression-free survival (PFS). In conclusion, we have developed a novel B7-H3 antibody that serves as a new tool for the detection of B7-H3 expression in AML and may help to facilitate risk stratification and treatment selection in AML patients.

B7-H3 enhances colorectal cancer progression by regulating HB-EGF via HIF-1α.

Background: B7-H3 (or CD276) represents an important costimulatory molecule expressed in many malignant solid tumors, including colorectal cancer (CRC). The receptor of B7-H3 is not known, and the intracellular function of B7-H3 remains obscure. Herein, we report that B7-H3 upregulated the epidermal growth factor heparin-binding epidermal growth factor (HB-EGF), likely by regulating hypoxia-inducible factor 1α (HIF-1α) and thereby promoting the progression of CRC. Methods: Lentiviral transfection was performed on CRC cells to establish stable low-B7-H3 expression cells. A mechanistic analysis with an Agilent human gene expression profiling chip was conducted on them. Clinical data and specimens were collected to detect the connection between B7-H3 and HB-EGF in CRC. Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to detect the messenger RNA (mRNA) level of B7-H3, HB-EGF, and HIF-1α. Chromatin immunoprecipitation (ChIP) quantitative real-time PCR was conducted. The protein level of HIF-1α and the phosphatidylinositide 3-kinases (PI3K)-protein kinase B (AKT) pathway were detected by western blot. HIF-1α was recovered by lentiviral transfection, and the HB-EGF mRNA levels, proliferation, invasion, and angiogenesis ability were detected. Results: B7-H3 promoted tumor progression through HB-EGF and the PI3K-AKT pathway. As B7-H3 was downregulated, HB-EGF levels were significantly reduced simultaneously, a growth trend that was shown by both CRC cell lines and cancer tissues. In addition, B7-H3 and HB-EGF had significant associations with tumor-node-metastasis (TNM) stage and lymph node metastasis in 50 CRC patients. The binding ability of HIF-1α to the HB-EGF promoter region was significantly decreased in the shB7-H3 RKO group. Western blot revealed that PI3K, AKT, and mammalian target of rapamycin (mTOR) protein amounts and p-AKT and p-mTOR phosphorylation were also downregulated in shB7-H3 RKO cells, suggesting that B7-H3 may regulate HIF-1α via PI3K-AKT signaling. After recovery of the HIF-1α level by lentiviral transfection, the HB-EGF mRNA levels, proliferation, invasion, and angiogenesis in CRC cells recovered as well. Conclusions: B7-H3 may transmit intracellular signals through PI3K-AKT-mTOR-HIF-1α signaling, upregulating HB-EGF. As the final transcription factor of the pathway, HIF-1α regulates the transcription of the HB-EGF gene, thereby promoting HB-EGF expression, which eventually mediates cell proliferation, invasion, and angiogenesis and promotes the progression of CRC.