Bispecific Antibody (BsAb) Design Service for Pretargeting System

Creative Biolabs is a prominent global provider in the expanding bispecific antibody (BsAb) market, with extensive expertise in designing and producing BsAbs for the pretargeting system.

Introduction

BsAbs are designed to bind two distinct targets, enabling targeted therapies. In pretargeting systems, BsAbs first bind to a specific target, such as a tumor antigen, and then a secondary agent is introduced to bind to the BsAb, enhancing targeted delivery. This approach improves treatment specificity, minimizes off-target effects, and is widely used in cancer immunotherapy, targeted drug delivery, and diagnostic imaging.

BsAb for Payload Delivery

BsAbs can be used for targeted payload delivery. The payload includes drugs, radiolabels, nanoparticles, or other small molecules. In radioimmunodetection, a key challenge is achieving high signal resolution over normal tissues, while in radioimmunotherapy, the focus is on delivering sufficient drugs to the target while sparing healthy tissues from toxicity. The pretargeting method separates the antibody's localization from the payload delivery, addressing these challenges. Unlike the 3-step method, BsAb-mediated delivery does not require blood-clearing agents, as BsAbs can naturally clear from the bloodstream. Initially, a cold BsAb binds to the target antigen, followed by a waiting period for clearance of unbound BsAbs. The payloads are administered after the unbound BsAbs have been removed from circulation. BsAbs can be employed to enhance payloads in tumor sites. This method greatly increases serum retention duration and improves the tumor/blood ratio.

Fig.1 The pretargeting approach steps and the most commonly applied pretargeting methods.^1,4

Pretargeting Radioimmunotherapy Based on BsAb

Pretargeting radioimmunotherapy using BsAbs is an innovative cancer treatment strategy. First, a BsAb targeting tumor-associated antigens is administered, localizing at the tumor site. Subsequently, a radiolabeled hapten, which binds specifically to the BsAb, is injected. This two-step approach minimizes systemic radiation exposure, improving the therapeutic index. BsAbs enable precise tumor targeting, while the rapid clearance of unbound radiolabeled hapten reduces off-target effects.

BsAb-Hapten Pretargeting Platform

BsAb-hapten pretargeting platforms have been widely studied, involving the use of BsAbs in combination with hapten-based radioligands. Two different strategies for BsAb-hapten pretargeting approaches have been studied clinically. Initially, fragmented bispecific antibody constructs (Fab-Fab'-chelate hapten) are mostly used, as this approach allows for shorter dosing intervals between the targeting construct and the radioligand, reducing the potential for hematologic toxicity. Over time, the technology advanced to the use of bispecific trivalent antibody constructs (Tri-Fab) in tandem with histamine-succinyl-glycine (HSG) peptide-based hapten radioligands. The Tri-Fab-HSG-hapten strategy offers high specificity and sensitivity for pretargeted PET imaging in cancer patients. Peptide-based haptens provide flexibility with radionuclides, maintaining stable antibody binding with lower risk.

Fig.2 Schematic overview of the pretargeting strategy.^2,4

A typical example of the Tri-Fab-HSG-hapten approach is the humanized Tri-Fab BsAb targeting carcinoembryonic antigen (CEA), which has been tested for pretargeted PET imaging and radioimmunotherapy in patients with colorectal cancer, medullary thyroid carcinoma, HER2-negative breast cancer, and metastatic lung cancer. Results showed higher sensitivity and specificity in detecting tumor lesions in metastatic colorectal cancer patients compared to ¹⁸F-FDG PET. It also effectively detected lesions in HER2-negative metastatic breast cancer patients.

Technologies of BsAbs Development for Pretargeting

Quadroma technology, a traditional method, fuses two antibody-producing hybridoma cell lines. Random heavy and light chain pairing yields a mixture of antibodies, requiring extensive screening to isolate the desired bispecific IgG with dual specificities (anti-TAA and anti-hapten). Its advantage lies in producing conventional IgG-like BsAbs, potentially retaining Fc-mediated effector functions.

Published Data

Researchers presented a versatile pretargeting platform for PET imaging using a novel 'two-step' immuno-PET approach with BsAbs. These BsAbs were designed for dual high-affinity binding to a fluorescein-based ¹⁸F-PET probe and tumor markers. A copper(I)-catalyzed click reaction was established for the radiosynthesis of [¹⁸F]TPF. The [¹⁸F]TPF was confirmed to bind to FITC-bearing BsAbs. The in vitro autoradiography evaluation revealed selective imaging of EpCAM-expressing OVCAR3 cells with the EpCAMxFITC BsAb. The platform's versatility was further shown with BsAbs targeting various cancer markers, including EpCAM, EGFR, MCSP, and PD-L1, indicating broad potential for future applications.

Fig.3 Autoradiography evaluation of the ability of cancer cell-bound BsAb EpCAMxFITC to capture [¹⁸F]TPF.^{3, 4}

Based on our advanced antibody engineering platform, Creative Biolabs offers comprehensive, one-stop BsAb services. Our BsAbs are rigorously validated and optimized for seamless integration with haptens in pretargeting systems to enhance the precision and effectiveness of your research project.