Dendritic cells (DCs) comprise diverse cell populations that play critical roles in antigen presentation and triggering immune responses in the body. However, several factors impair the immune function of DCs and may promote immune evasion in cancer. Understanding the mechanism of DC dysfunction and the diverse functions of heterogeneous DCs in the tumor microenvironment (TME) is critical for designing effective strategies for cancer immunotherapy. Clinical applications targeting DCs summarized in this report aim to improve immune infiltration and enhance the biological function of DCs to modulate the TME to prevent cancer cells from evading the immune system. Herein, factors in the TME that induce DC dysfunction, such as cytokines, hypoxic environment, tumor exosomes and metabolites, and co‐inhibitory molecules, have been described. Furthermore, several key signaling pathways involved in DC dysfunction and signal‐relevant drugs evaluated in clinical trials were identified. Finally, this review provides an overview of current clinical immunotherapies targeting DCs, especially therapies with proven clinical outcomes, and explores future developments in DC immunotherapies.

Background: Triple-negative breast cancer (TNBC) is more prone to distant metastasis and visceral recurrence in comparison to other breast cancer subtypes, and is related to dismal prognosis. Nevertheless, TNBC has an undesirable response to targeted therapies. Therefore, to tackle the huge challenges in the diagnosis and treatment of TNBC, Nectin-4 was selected as a theranostic target because it was recently found to be highly expressed in TNBC. We developed anti-Nectin-4 monoclonal antibody (mAb_Nectin-4)-based theranostic pair, ^99mTc-HYNIC-mAb_Nectin-4 and mAb_Nectin-4-ICG. ^99mTc-HYNIC-mAb_Nectin-4 was applied to conduct immuno-single photon emission computed tomography (SPECT) for TNBC diagnosis and classification, and mAb_Nectin-4-ICG to mediate photothermal therapy (PTT) for relieving TNBC tumor growth. Methods: Nectin-4 expression levels of breast cancer cells (MDA-MB-468: TNBC cells; and MCF-7, non-TNBC cells) were proved by western blot, flow cytometry, and immunofluorescence imagning. Cell uptake assays, SPECT imaging, and biodistribution were performed to evaluate Nectin-4 targeting of ^99mTc-HYNIC-mAb_Nectin-4. A photothermal agent (PTA) mAb_Nectin-4-ICG was generated and characterized. In vitro photothermal therapy (PTT) mediated by mAb_Nectin-4-ICG was conducted under an 808 nm laser. Fluorescence (FL) imaging was performed for mAb_Nectin-4-ICG mapping in vivo. In vivo PTT treatment effects on TNBC tumors and corresponding systematic toxicity were evaluated. Results: Nectin-4 is overexpressed in MDA-MB-468 TNBC cells, which could specifically uptake ^99mTc-HYNIC-mAb_Nectin-4 with high targeting in vitro. The corresponding immunoSPECT imaging demonstrated exceptional performance in TNBC diagnosis and molecular classification. mAb_Nectin-4-ICG exhibited favourable biocompatibility, photothermal effects, and Nectin-4 targeting. FL imaging mapped biodistribution of mAb_Nectin-4-ICG with excellent tumor-targeting and retention in vivo. Moreover, mAb_Nectin-4-ICG-mediated PTT provided advanced TNBC tumor destruction efficiency with low systematic toxicity. Conclusion: mAb_Nectin-4-based radioimmunoimaging provides visualization tools for the stratification and diagnosis for TNBC, and the corresponding mAb_Nectin-4-mediated PTT shows a powerful anti-tumor effect. Our findings demonstrate that this Nectin-4 targeting strategy offers a simple theranostic platform for TNBC.

Many methods have been developed to produce bispecific antibodies (BsAbs) for industrial application. However, huge challenges still remain in synthesizing whole length BsAbs, including their assembly, stability, immunogenicity, and pharmacodynamics. Here we present for first time a generic technology platform of generating bispecific IgG antibodies, “Bispecific Antibody by Protein Trans-splicing (BAPTS)”. Different from published methods, we assembled two parental antibody fragments in the hinge region by the protein trans-splicing reaction of a split intein to generate BsAbs without heavy/heavy and light/heavy chain mispairing. Utilizing this simple and efficient approach, there have been several BsAbs (CD3×HER2, CD3×EGFR, EGFR×HER2) synthesized to demonstrate its broad applicability. Correctly paired mAb arms were assembled to form BsAbs that were purified through protein A affinity chromatography to demonstrate industrial applicability at large scale. Further, the products were characterized through physical-biochemistry properties and biological activities to confirm expected quality of the products from “BAPTS”. More importantly, correct pairing was confirmed by mass spectrum. Proof-of-concept studies with CD3×HER2 BsAb (T-cell recruitment) demonstrated superior bioactivity compared with trastuzumab. The results of undetectable mispairing and high biological activity have indicated that this method has the potential to be utilized to manufacture BsAbs with high efficiency at industrial scale.

Recombinant human interleukin-15 (IL-15) is a potent cancer immunotherapeutic candidate due to its excellent immune stimulating effects. Previous work demonstrated that IL-15 appeared with short half-life in circulation system, while the complex with its receptor can prolong the half-life as well as benefit its activities in vivo. Therefore, IL-15 complex was more favorably considered for clinical development. Herein we developed IL-15·sIL-15Rα/Fc, a complex comprising of IL-15 and the extracellular region of its receptor alpha subunit which fused to Immunoglobulin G (IgG1) Fc to further prolong the half-life in plasma. Through transient gene expression in HEK293 cells, we expressed the superagonist by co-transfection of plasmids encoding IL-15 and sIL-15Rα/Fc respectively, yielding 36 mg/L of product after purification. Pharmacokinetic study demonstrated that the combination profoundly prolonged the half-life of IL-15 to 13.1 h in mice, about 18 folds longer than that of IL-15 monomer which is around 0.7 h. The bioactivity of the superagonist was characterized by CTLL-2 cells proliferation assay in vitro, showing its capability of stimulating the expansion of memory CD8⁺ T cells (cluster of differentiation) in mouse spleen. Using a HT-29 xenograft NOD-SCID mouse model, we observed tumor growth inhibition in all groups that received the superagonist, indicating its anti-tumor efficacy via stimulating infused human immune cells. In addition, combo cancer treatment by IL-15·sIL-15Rα/Fc and programmed death-1 (PD-1) antibody have shown stronger inhibitory effects as compared with treatment with either single molecule. Therefore, we developed IL-15·sIL-15Rα/Fc to be a long half-life potential cancer immunotherapy candidate that can be applied alone or in synergy with PD-1/PD-L1 blockade.

Background: Prolactin receptor (PRLR) is highly expressed in a subset of human breast cancer and prostate cancer, which makes it a potential target for cancer treatment. In clinical trials, the blockade of PRLR was shown to be safe but with poor efficacy. It is therefore urgent to develop new therapies against PRLR target. Bispecific antibodies (BsAbs) could guide immune cells toward tumor cells, and produced remarkable effects in some cancers.Methods: In this study, a bispecific antibody targeting both tumor antigen PRLR and T cell surface CD3 antigen (PRLR-DbsAb) was constructed by split intein mediated protein transsplicing (BAPTS) system for the first time. Its binding activity was determined by Biacore and Flow cytometry, and target-dependent T cell mediated cytotoxicity was detected using LDH release assay. ELISA was utilized to study the secretion of cytokines by immune cells. Subcutaneous tumor mouse models were used to analyze the in vivo anti-tumor effects of PRLR-DbsAb.Results: PRLR-DbsAb in vitro could recruit and activate T cells to promote the release of Th1 cytokines IFN-γand TNF-α, which could kill PRLR expressed breast cancer cells. In xenograft models with breast cancer cell line T47D, NOD/SCID mice intraperitoneally injected with PRLR-DbsAb exhibited significant inhibition of tumor growth and a longer survival compared to mice treated with PRLR monoclonal antibody (PRLR mAb).Conclusions: Both in vitro and in vivo experiments showed PRLR-DbsAb had a potential therapy of cancer treatment potential therapy for cancer. Immunotherapy may be a promising treatment against the tumor target of PRLR.