Multiplex Immunofluorescence: A Powerful Tool in Cancer Immunotherapy.

Traditional immunohistochemistry (IHC) has already become an essential method of diagnosis and therapy in cancer management. However, this antibody-based technique is limited to detecting a single marker per tissue section. Since immunotherapy has revolutionized the antineoplastic therapy, developing new immunohistochemistry strategies to detect multiple markers simultaneously to better understand tumor environment and predict or assess response to immunotherapy is necessary and urgent. Multiplex immunohistochemistry (mIHC)/multiplex immunofluorescence (mIF), such as multiplex chromogenic IHC and multiplex fluorescent immunohistochemistry (mfIHC), is a new and emerging technology to label multiple biomarkers in a single pathological section. The mfIHC shows a higher performance in cancer immunotherapy. This review summarizes the technologies, which are applied for mfIHC, and discusses how they are employed for immunotherapy research.

Near Infrared Photoimmunotherapy: A Review of Recent Progress and Their Target Molecules for Cancer Therapy.

Near infrared photoimmunotherapy (NIR-PIT) is a newly developed molecular targeted cancer treatment, which selectively kills cancer cells or immune-regulatory cells and induces therapeutic host immune responses by administrating a cancer targeting moiety conjugated with IRdye700. The local exposure to near-infrared (NIR) light causes a photo-induced ligand release reaction, which causes damage to the target cell, resulting in immunogenic cell death (ICD) with little or no side effect to the surrounding normal cells. Moreover, NIR-PIT can generate an immune response in distant metastases and inhibit further cancer attack by combing cancer cells targeting NIR-PIT and immune regulatory cells targeting NIR-PIT or other cancer treatment modalities. Several recent improvements in NIR-PIT have been explored such as catheter-driven NIR light delivery, real-time monitoring of cancer, and the development of new target molecule, leading to NIR-PIT being considered as a promising cancer therapy. In this review, we discuss the progress of NIR-PIT, their mechanism and design strategies for cancer treatment. Furthermore, the overall possible targeting molecules for NIR-PIT with their application for cancer treatment are briefly summarised.

EpCAM- and EGFR-Specific Antibody Drug Conjugates for Triple-Negative Breast Cancer Treatment.

Triple-negative breast cancer (TNBC) is a group of heterogeneous and refractory breast cancers with the absence of estrogen receptor (ER), progesterone receptor (PgR) and epidermal growth factor receptor 2 (HER2). Over the past decade, antibody drug conjugates (ADCs) have ushered in a new era of targeting therapy. Since the epidermal growth factor receptor (EGFR) and epithelial cell adhesion molecule (EpCAM) are over expressed on triple-negative breast cancer, we developed novel ADCs by conjugating benzylguanine (BG)-modified monomethyl auristatin E (MMAE) to EpCAM- and EGFR-specific SNAP-tagged single chain antibody fragments (scFvs). Rapid and efficient conjugation was achieved by SNAP-tag technology. The binding and internalization properties of scFv-SNAP fusion proteins were confirmed by flow cytometry and fluorescence microscopy. The dose-dependent cytotoxicity was evaluated in cell lines expressing different levels of EGFR and EpCAM. Both ADCs showed specific cytotoxicity to EGFR or EpCAM positive cell lines via inducing apoptosis at a nanomolar concentration. Our study demonstrated that EGFR specific scFv-425-SNAP-BG-MMAE and EpCAM-specific scFv-EpCAM-SNAP-BG-MMAE could be promising ADCs for the treatment of TNBC.

Toward Homogenous Antibody Drug Conjugates Using Enzyme-Based Conjugation Approaches.

In the last few decades, antibody-based diagnostic and therapeutic applications have been well established in medicine and have revolutionized cancer managements by improving tumor detection and treatment. Antibodies are unique medical elements due to their powerful properties of being able to recognize specific antigens and their therapeutic mechanisms such as blocking specific pathways, antibody-dependent cellular cytotoxicity, and complement-dependent cytotoxicity. Furthermore, modification techniques have paved the way for improving antibody properties and to develop new classes of antibody-conjugate-based diagnostic and therapeutic agents. These techniques allow arming antibodies with various effector molecules. However, these techniques are utilizing the most frequently used amino acid residues for bioconjugation, such as cysteine and lysine. These bioconjugation approaches generate heterogeneous products with different functional and safety profiles. This is mainly due to the abundance of lysine and cysteine side chains. To overcome these limitations, different site-direct conjugation methods have been applied to arm the antibodies with therapeutic or diagnostics molecules to generate unified antibody conjugates with tailored properties. This review summarizes some of the enzyme-based site-specific conjugation approaches.